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Notes: Dielectric relaxation peak due to bound water was found around 100 MHz in poly(dG-dC) · poly(dG-dC) and calf thymus DNA in water-ethanol mixtures with NaCl buffer. Relaxation time and strength show a transition for poly(dG-dC) · poly(dG-dC) at anethanol composition Cw = 0.45 (w/w) where the structural transition from B- to Z-DNA takes place. It has been suggested that the transition is caused by removal of the bound water molecules preferentially from the phosphate groups. If the bound water molecules are removed equally from the phosphate groups and the grooves, the structural transition from B to A takes place. By analogy with hydration of tropocollagen, it was found that 19 water molecules per one nucleotide are at least necessary to keep B-DNA. Thirteen molecules are bound to A-DNA and 9 molecules to Z-DNA. Stringlike multimers are proposed as available structures of the bound water.

Notes: A simple bead model is proposed for the antibody molecule immunoglobulin IgG1. The partial flexibility of the hinge is represented by a quadratic potential associated to the angles between arms. Conformational and hydrodynamic properties are calculated using Monte Carlo (rigid-body) and Brownian dynamics simulations. Comparison of experimental and calculated values for some overall properties allows the assignment of dimensions and other model parameters. The Brownian dynamics technique is used next to simulate a rotational correlation function that is comparable with the decay of fluorescence emission anisotropy. This is done with varying flexibility at the hinge. The longest relaxation time shows a threefold decrease when going from the rigid Y-shaped conformation to the completely flexible case. The calculations are in good agreement with the decay times observed for IgG1. A flexibility analysis of the latter indicates that a variability of ±55° (standard deviation) in the angle between the Fab arms.

Notes: Computational techniques have been used to aid interpretation of observed systematic shifts in the amide III frequencies of Ala-X peptides. Optimized structures and frequencies have been calculated for Ala-X peptides using GAUSSIAN86/88 with the 4-31G basis, MOPAC, and normal mode methods based on empirical force fields. We observe the following: (1) Frequencies calculated using scaled GAUSSIAN86 force constants correlate well with the experimental results. (2) Structures of the Ala-X peptides optimized by GAUSSIAN show a clear trend toward lower values of the dihedral angle φ as the X side chain becomes larger, while structures optimized here using semiempirical and empirical force fields do not show trends. (3) Computational changes in peptide conformations from β-sheet to α-helix produce large changes in both amide I and amide III frequencies that are inconsistent with the experimental results. (4) Computational changes in the dihedral angle φ of Ala-Ala produce a change in the amide III frequency consistent with the experimental results. (5) The experimental frequency shifts cannot be attributed directly to the effects of changing mass.

Notes: Photon correlation spectroscopy was used to study both F-actin and F-actin/filamin networks in solution. The measured autocorrelation functions were analyzed with the inverse Laplace transform CONTIN. The resulting frequency distributions consist of maximal five relatively narrow peaks. This rather unexpected finding disagrees with the frequency spectra calculated for an entangled rigid rod model. For this model, we expect spectra consisting of a single broad peak. Factors like flexibility, deviation from an experimental length distribution, interference with chemical reactions, and translation rotation coupling that would influence the profile of the frequency distribution could be excluded by comparison of the data with model calculations or qualitative estimates. We conclude that our data for F-actin are consistent with the dynamics of an infinite network stable on the time scale of the observed modes. Further support for this interpretation is provided by a comparison with frequency spectra of actin/filamin networks. These appear to be very similar in shape. The main peak of these spectra that corresponds to the slow motions shifts to lower frequencies with increasing cross-link density. It appears that higher frequency modes of the gel are more efficiently damped with progressive cross-link density, resulting in a predominance of the slow motions in the spectra. This behavior has been already found in other systems and seems to be a general feature of cross-linked systems.

Notes: The isomeric decapeptides Boc-Aib-Ala-Leu-Ala-Aib-Aib-Leu-Ala-Leu-Aib-OMe (II) and Boc-Aib-Ala-Aib-Ala-Leu-Ala-Leu-Aib-Leu-Aib-OMe (III), are predominantly a-helical with little effect on the conformation with interchange of Aib/Ala residues or Aib/Leu residues. The packing motif of helices in crystal II is antiparallel, whereas the helices pack in a skewed fashion in crystal III, with a 40° angle between neighboring helix axes. Crystal III contains a water molecule in a hydrophobic hole that forms hydrogen bonds with two carbonyl oxygens that also participate in 5 → 1 type hydrogen bonds. Values for helical torsional angles φ and ψ assume a much wider range than anticipated. Crystal II: C49H88N10O13, space group P21 with a = 16.625(2) Å, b = 9.811(5) Å, c = 18.412(3) Å, β = 99.79(1)°, Z = 2, R = 5.7% for 4338 data with |F0 | 〉 3σ(F). Crystal III: C49H88N10O13. 1/2H2O, space group P21, with a = 11.072(2) Å, b = 34.663(5) Å, c = 16.446(3) Å, β = 107.85(1) °, Z = 4, R = 8.3% for 6087 data with |F0| 〉 3σ(F).

Notes: The conformation of Boc-Pro-Ser-NHCH3 (1) was studied in the solid state and in solution. In the crystal, the steric structure of 1 is characterized by an E(cis) urethane tertiary amide bond and the lack of intramolecular H bonds. Four-hundred megahertz 1H- and 101-MHz 13C-nmr studies in CDCl3 clearly show the presence of two conformers differing in the rotameric state of the tertiary-amide bond. Selective 1H-13C nuclear Overhauser enhancement experiments at -20°C as well as 1H-nmr and ir data indicate that the major trans conformer (84% in CDC13) may adopt a type I β-turn conformation with a possible Oγ - H-N interaction, similarly to Piv-Pro-Ser HCH3 (2) [A. Aubry, N. Ghermani, and M. Mar-raud (1984) Int. J. Peptide Protein Res. 23, 113-122]. Molecular mechanics calculations on the cis rotamer show that the β-pleated-like backbone conformation of serine in the crystal of 1 is not a low-energy state for the isolated molecule; it is caused by packing forces, particularly by the helical network of intermolecular H bonds.

Notes: Poly [d(Gm5C)·d(Gm5C)] can be titrated from the Z form in 30% ethanol to the Z′ form by adding either ethanol or divalents. Analysis by singular-value decomposition of CD spectra recorded during the titration reveal that ethanol and transition metal induced changes are two state with a single-step titration curve. When the change is induced by arkaline earth metals it is still two state, but in contrast the titration curve is complex, indicating two binding sites.

Notes: β-Endorphin has been studies iin SDS micelles by one- and two-dimensional nmr spectroscopy (1D and 2D nmr), and to explore the influence of peptide length and composition on the polypeptide structure, the investigation was extended to a number of fragments. The nmr results are compared with those obtained from CD experiments and discussed in terms of a secondary structure that involves the central region of β-endorphin.

Notes: Nonpolymerizable tropomyosin (NPTm) is found to unfold thermally at high ionic strength almost exactly as the parent protein, but it does not aggregate at low ionic strength. Thus, NPTm can be used as a tropomyosin surrogate whose coiled-coil structural stability can be probed by varying the ionic strength. Studies of NPTm by CD show that increasing ionic strength stabilizes the coiled-coil structure. CD spectra over a wide range of helix content, obtained by varying either temperature or ionic strength, show an isodichroic point at 203 nm, suggesting a local, residue-level, two-state model. At given temperature, such a local helix ⇆ random equilibrium suggests In [Φh/(l - Φh)] = A1 + A2In, wherein Φh is the fraction helix, and A1, A2 and n are constants. In the low ionic strength region, theoretical limiting laws for ionic strength mediated charge-charge, dipole-dipole, and apolar-apolar (salting out) interactions give, respectively, n = 0.5, 1.0, and 1.0. Our experimental values for 40°C, where the data span a wide range of helix content, show n = 1.0, suggesting that ionic strength stabilizes either by reducing dipole-dipole repulsions or by enhancing hydrophobic interactions, both probably interhelix in nature. Two segments of tropomyosin, 11Tm127 and 142Tm281, neither of which aggregate at low ionic strength, give results similar to those for NPTm, i.e., n = 0.96 and 0.84, respectively.

Notes: The L-azetidine-2-carboxylic acid (Aze) residue can be incorporated into proteins in the place of L-proline, of which it is the lower homologue. This substitution alters the properties of proteins, especially of collagen. Conformational constraints in N-acetyl-Aze-N′-methylamide and in several dipeptides containing Aze have been analyzed by means of energy computations. They have been compared with peptides containing Pro. The overall con-formational preferences of Aze and Pro are similar, but several significant differences occur between them. In general, peptides containing Aze are somewhat more flexible than corresponding peptides containing Pro, because of a decrease in constraints caused by repulsive nonconvalent interactions of the atoms of the ring with neighboring residues. This results in an entropic effect that lessens the stability of ordered polypeptide conformations with respect to the disordered statistical coil. The collagen-like near-extended conformation is energetically less favorable for Aze than for Pro in the single residue and in dipeptides. This effect also contributes to a destabilization of the collagen triple helix. The influence of Aze on the conformation of polypeptides is discussed in the accompanying papers.

Notes: According to counterion condensation theory, one of the contributions to the polyelectrolyte free energy is a pairwise sum of Debye-Hückel potentials between polymer charges that are reduced by condensed counterions. When the polyion model is taken as an infinitely long and uniformly spaced line of charges, a simple closed expression for the summation, combined with entropy-derived mixing contributions, leads to the central result of the theory, a condensed fraction of counterions dependent only on the linear charge density of the polyion and the valence of the counterion, stable against increases of salt up to concentrations in excess of 0.1M. Here we evaluate the sum numerically for B-DNA models other than the infinite line of B-DNA charges. For a finite-length line there are end effects at low salt. The condensation limit is reached as a flat plateau by increasing the salt concentration. At a fixed salt concentration the condensation limit is reached by increasing the length of the line. At moderate salt even very short B-DNA line-model oligomers have condensed fractions not far from the infinite polymer limit. For a long double-helical array with charge coordinates at the phosphates of B-DNA, the limiting condensed fraction appears to be approached at low salt. In contrast to the results for the line of charges, however, the computed condensed fraction varies strongly with salt in the range of experimentally typical concentrations. Salt invariance is restored, in agreement with both the line model and experimental data, when dielectric saturation is considered by means of a distance-dependent dielectric function. For sufficiently long B-DNA line and helical models, at typical salt concentrations, the counterion binding fraction approaches the polymer limit as a linear function of 1/P, where P is the number of phosphate groups of B-DNA.

Notes: Monte Carlo computer simulation is described for the dinucleotide duplex rGpC together with 562 water molecules at an environmental density of 1 g/cc in a cubic cell under periodic boundary conditions. Water-water interactions were treated using the TIP4P potential and the solute water interactions by TIP4P spliced with the nonbonded interactions from the AMBER 3.0 force field. The simulation was subjected to proximity analysis to obtain solute coordinate numbers and pair interaction energies for each solute atom. Hydration density distributions partitioned into contributions from the major groove side, the minor groove side and the sugar-phosphate backbone were examined, and the probabilities of occurrence for one- and two-water bridges in the simulation were enumerated. The results were compared with observations of crystallographic ordered water sites from x-ray diffraction studies on G and C containing small molecules, and in crystal structure determinations of the sodium, calcium, and ammonium salts of rGpC. The calculated results are generally consistent with the observed sites, except for cytosine N4, where a hydration site is predicted yet none observed in rGpC salts, and for guanine N3, which appears in this calculation to compete unfavorably with the adjacent donor site at guanine N2. There is, however, a significant probability of finding a one-water G-N3-W-G-N2 bridge indicated in the simulation. An explanation for the guanine N3 discrepancy in terms of electrostatic potentials is also offered. The calculated one-and two-water bridges in the rGpC hydration complex coincide in a number of cases to those observed in the ordered water structure of the sodium rGpC crystal hydrate.

Notes: The energy of formation of salt linkages between Arg or Lys with Asp or Glu in a polypeptide chain having the collagen fold have been estimated using the fully empirical energy minimization scheme AMBER. The polypeptide was considered both in an isolated and a hydrated triple helical state. The collagen fold associated with a one-bonded triple helical conformation allows intrachain salt linkages having stabilization energies of 60-100 kcal when the reacting residues are separated by no more than two intervening residues. The amino end of one side chain always approaches the carboxyl end of the other side chain, and simultaneously approaches the carbonyl oxygen of the intervening backbone residue. The salt linkage conformation and the backbone conformation of the isolated collagen fold in vacuo are maintained when the molecules are in a hydrated triple helix. These results are compatible with a fold-forming role for salt linkages, especially in proline poor regions, during collagen polypeptide synthesis, and with the persistence of intrachain salt linkages throughout molecular and fibril assembly.

Notes: Molecular dynamics simulations were carried out without explicit consideration of solvent to explore the conformational mobility of blood group A and H oligosaccharides. The potential energy force field of Rasmussen and co-workers was used with the CHARMM program on a number of disaccharide and trisaccharide models composed of fucose, galactose, glucose, N-acetyl glucosamine, and N-acetyl galactosamine chosen to represent various fragments of blood group oligosaccharides. In agreement with results of earlier studies, stable chair conformations were found for each pyranoside from which no transitions were detected in simulations as long as 800 ps. Exocyclic dihedral angles, including that of C5 - C6, generally show numerous transitions on a time scale of approximately 5-30 ps. The dihedral angles of some but not all glycosidic linkages of blood group oligosaccharides show transitions on the time scale of 30-50 ps, implying that the extent of internal motion in blood group oligosaccharides depends strongly on linkage stereochemistry. For certain blood group A and H oligosaccharides that show limited internal motion in these simulations, we argue that the calculations are consistent with our previous analysis of 1H nuclear Overhauser enhancement (NOE) data that imply single conformations over a wide range of temperature and solvent conditions. While the trajectories are consistent with 13C T1 values that have been interpreted as indicating rigid conformations, measurements of 13C-NOE and T1 as a function of magnetic field strength are proposed as an improved method for experimental detection of the internal motion that is suggested for certain oligosaccharides in these simulations. The results of these simulations differ substantially from those of peptides of a similar molecular weight in that the oligosaccharides show much less internal motion.

Notes: A search procedure is described for making stereospecific assignments at prochiral centers in proteins on the basis of nuclear Overhauser enhancement and coupling constant data derived from nmr experiments. A data base comprising torsion angles, associated 1H-1H coupling constants and interproton distances is searched by a computer algorithm for sets of values that match the experimental data within specified error limits. Two different data bases are used. The first is a crystallographic data base derived from 34 well-refined crystal structures; the second is a systematic data base derived from conformations of a short peptide fragment with idealized geometry by systematically varying the φ,ψ, and χ1 torsion angles. Both approaches are tested for β-methylene groups with model data obtained from 20 crystal structures. The results for the two methods are similar though not identical, so that a combination of the two methods appears to be useful. With an appropriate choice of error estimates, around 80% of the β-methylene groups could be assigned in the test calculations. In addition, results with experimental nmr data indicate that a similar percentage of stereospecific assignments can be made in practical situations.

Notes: Two extant models of thermal folding/unfolding equilibria in two-chain, α-helical coiled coils are tested by comparison with experimental results on excised, isolated subsequences of rabbit αα-tropomyosin (Tm). These substances are designated iTmj where i and j are, respectively, the residue numbers (in the 284-residue parent chain) of the N- and C-terminal residues of the subsequence. One model postulates that a coiled coil consists of segments, each denaturing in an all-or-none manner, like small globular proteins. Thus this model yields a small number of populated molecular species. In an extant calorimetry study of 11Tm127 and of 190Tm284, each required only two all-or-none-segments, and their enthalpies and transition temperatures were assigned. These assignments are shown here to yield the concentration of all molecular species, and therefore the helix content, as a function of temperature. Such calculations for 190Tm284 are in tolerable agreement with CD experiments, but those for 11Tm127 are in gross disagreement. Thus, either the model itself or the calorimetric assignment is faulty. In the second model, all conformational states are counted and weighted, as in the Zimm-Bragg theory for single-chain polypeptides. This theory has been extended (by Skolnick) to two-chain coiled coils and is here used to fit CD data for 11Tm127, 142Tm281, and 190Tm284. The fit is tolerable for 11Tm127, good for 142Tm281, and quantitative for 190Tm284. Thus this comparison does not falsify this second model. The helix-helix interaction free energy, obtainable from the fit, shows nonadditivity when isolated subsequences are compared with the parent. This suggests that removal of a region from a long coiled coil allows energetically substantial adjustments in side-chain packing in the helix-helix interface. Thus, the helix-helix interaction in long coiled coils is characteristic of a global free energy minimum and not just of the regional constellation of side chains.

Notes: The effect of elevated hydrostatic pressure on the secondary structure of poly(L-lysine) was studied using Fourier transform ir spectroscopy. According to changes observed in the amide I band, both the β-sheet and the unordered polypeptide undergo a reversible, pressure-induced conformational change to α-helix. The conversion occurs at a much higher pressure from the unordered conformer (∼ 9 kbar) than from the β-sheets (∼ 2 kbar). The structural changes were found to be slower at pH 〉 11, especially at the highest concentration investigated (10 wt%), reflecting the fact that extensive hydrogen-bond networks have to reorganize. This study shows that alterations of polypeptidic conformations induced by elevated hydrostatic pressure are reversible, but that an apparent irreversibility can result from kinetic factors in the case of conformational changes involving extensive rearrangements. The present results also show that the strength of the hydrogen bonds between the backbone amide groups is not the only factor that determines the closest packing of the polypeptide molecules.

Notes: The synthesis of some repetitive sequences of elastin and their simplified analogues, all comprising the structural unit Gly-X-Gly (X = Val, Leu, Ala), is described. In particular, the following peptides and polypeptides were synthesized and characterized: Boc-Gly-Val-Gly-Gly-Leu-OMe, Boc-Gly-Leu-Gly-Gly-Val-OMe, Boc-(Gly-Val-Gly-Gly-Leu)2-OMe, Boc-(Gly-Val-Gly-Gly-Leu)3-OMe, Boc-Gly-Val-Gly-Gly-OEt, Boc-Leu-Gly-Gly-Leu-OMe, Boc-Val-Gly-Gly-Val-OMe, poly(Ala-Gly-Gly), poly(Val-Gly-Gly), and poly (Leu-Gly-Gly).In every case, the synthesis was accomplished by classical procedures in solution, by using the p-nitrophenyl ester method for the polycondensation step, and the mixed anhydride or the azide methods for the coupling steps.

Notes: Conformational studies on synthetic repetitive sequences and analogues of elastin are described.CD and nmr measurements gave evidence of flexible β-turns as the dominant structural feature whose stability was found to decrease by increasing the number of repetitive units. The sequences comprised the structural unit Gly-X-Gly (X = Val, Leu, Ala), with X-Gly or Gly-Gly located at the corners of the bend.Based on that, it is proposed that these regions of elastin, unlike the proline-containing sequences, contribute to the elasticity of the protein through a classical mechanism in terms of the rotational isomeric state theory.

Notes: The indirect solvent-induced effect on the free energy of binding of biopolymers is examined within the framework of classical statistical mechanics. We focus specifically on the role of the solute-solvent hydrogen bonding. In particular, we have estimated the first order solvent effedt on the indirect interaction between two biopolymers. We find that the solvent-induced interactios between two hydrophilic groups through water-bridged hydrogen bonds could significantly enhance the binding free energy. Some preliminary estimates indicate that this effect is significant and perhaps could be crucial in molecular recognition processes. Furthermore, we have calculated, from crystal structure data, the distance distribution between all the oxygens and nitrogens on the surface of some proteins that do not belong to the binding domain. In most cases we found an enhanced peak in the range of 4-5 Å, which is where we expect to find strong solvent-induced interactions.

Notes: The translational and rotational diffusion coefficients and the intrinsic viscosity of semiflexible, randomly broken, and wormilike chains have been obtained by Monte Carlo simulation in the context of the rigid-body treatment. Both approximate and rigorous rigid-body hydrodynamics are used, so that the error introduced by the approximate methods can be evaluated. A randomly broken chain and a wormilike chain having the same contour length and persistence length have the same radius of gyration but different values for any of the hydrodynamic properties. The two types of chains are compared in this regard. Considering that the cross section of the chain is represented by a cylinder better than by a string of spheres, we devise a cylindrical correction to be applied to the results simulated for chains of beads. Application is made to the analysis of experimental data for the translational and rotational coefficients of DNA fragments with up to 103 base pairs, obtaining the persistence length for each model. The values for the wormlike chain agree well with model-independent values obtained from radii of gyration and with other literature data at varying ionic strength. The randomly broken chain is equally able to reproduce the experimental length dependence of the properties, but the resulting persistence length may be too high.

Notes: Quaterpyridineiron(III) complex ions (FeT) anchored to ordered poly(L-glutamate) or poly(D-glutamate) were used as enantiomeric systems for stereoselective oxidation of L-cysteine, a widely encountered ligand in both heme and nonheme iron-sulfur proteins. Surprisingly, electron transfer from substrate to the central metal ion does not take place, despite the favorable thermodyamic driving force. Instead, a stable, polymer-supported, Fe(III)T-cysteinate complex forms, to which no stereoselectivity is associated. This finding contrasts the known lability of iron(III)-thiolate complexes, ultimately yielding disulfides and iron(II) species, and is relevant in view of the current interest in model compunds of cytochrome P-450, where cysteine is axially bound to ferric prophyrin. The formation of this complex was studied by kinetics and low-temperature electron paramagnetic resonance spectroscopy, while the stereochemical features of the diastereomeric encounter intermediates were investigated by theoretical conformational analysis. The results collectively emphasize the environmental contraol of the polypeptide matrix on the stability of cysteinate sulfur axial ligation. Implications of the absence of stereoselectivity in the reaction investigated are also discussed in the light of a few general considerations concerning chiral discrimination in reactions between optically active compounds.

Notes: Pyridoxylated adult human hemoglobin (HbAo) was prepared using a one molar equivalent of pyridoxal 5-phosphate (PLP) per heme and reduced with either NaCNBH3 or NaBH4. A separate sample was pyridoxylated and passed through a mixed-bed ion exchange column without reduction. All three preparations had a P50 of 29 ± 2 torr and a cooperativity of n = 2.4 ± 0.1. These preparations, in both the oxy and deoxy forms, were then treated with 7 equivalents of glutaraldehyde per tetramer at pH 6.8 at 4°C and at room temperature. The polymerization invariably reduced the P50 to 18 ± 2 torr with Hill coefficients of less than 2. These solutions, with or without further reduction using NaCNBH3, all retained the PLP in differing amounts (2-3 moles/tetramer). Methemoglobin concentrations were increased during the polymerization reaction. The normal pyridoxylation procedure, using sodium borohydride reduction, resulted in a number of different molecular species. Polymerization with glutaraldehyde caused a further proliferation of molecular species that could not be separated by anion exchange chromatography or by isoelectric focusing. The extent of polymerization, estimated by gel exclusion chromatography and SDS polyacrylamide gel electrophoresis, was from 40 to 50%. Analysis of the reverse phase chromatograms, which separate the heme and the α- and β-chains, showed extensive polymerization and distribution of the radioactively labeled PLP on the protein for all preparations. All of the polymerized and pyridoxylated samples were unstable, and showed different chromatographic patterns after storage at 4°C for 1 month. Attempts to stabilize these preparations by further reduction with NaCNBH3 gave products with a lower P50 and lower cooperativity. When the reactions were conducted with a purified HbAo, heterogeneity was somewhat decreased compared to the normally used stroma-free hemoglobin, but a large number of molecular species were still formed.

Notes: The peptide N-Boc-L-Phe-dehydro-Leu-L-Val-OCH3 was synthesized by the usual workup procedure and finally by coupling the N-Boc-L-Phe-dehydro-Leu-OH to valine methyl ester. It was crystallized from its solution in methanol-water mixture at 4°C. The crystals belong to the triclinic space group P1 with a = 5.972(5) Å, b = 9.455(6) Å, c = 13.101(6) Å, α = 103.00(4)°, β = 97.14(5)°, γ = 102.86(50)°, V = 690.8(8) Å, Z = 1, dm = 1.179(5) Mg m-3 and dc = 1.177(5) Mg m-3. The structure was determined by direct methods using SHELXS86. It was refined by block-diagonal least-squares procedure to an R value of 0.060 for 1674 observed reflections. The C2α-C2β distance of 1.323(9) Å in dehydro-Leu is an appropriate double bond length. The bond angle Cα-Cβ-Cγ in the dehydro-Leu residue is 129.4(8)°. The peptide backbone torsion angles are θ1 = -168.6(6)°, ω0 = 170.0(6)°, φ1 = -44.5(9)°, ψ1 = 134.5(6)°, ω1 = 177.3(6)°, φ2 = 54.5(9)°, ψ2 = 31.1(10)°, ω2 = 171.7(6)°, φ3 = 51.9(8)°, ψ3T = 139.0(6)°, θT = -175.7(6)°. These values show that the backbone adopts a β-turn II conformation. As a result of β-turn, an intramolecular hydrogen bond is formed between the oxygen of the ith residue and NH of the (i + 3)th residue at a distance of 3.134(6) Å. The Boc group has a trans-trans conformation. The side-chain torsion angles of the Phe residue are χ1 = 171.6(6)°, χ12,1 = -102.1(9)°, and χ12,2 = 78.6(10)°. The side-chain conformational angles of dehydro-Leu residue are χ2 = 2.7(13)°, χ22,1 = -107.3(11)°, and χ22,2 = 131.3(10)°. The torsion angles χ31,1 and χ31,2 that define the conformation of the valyl side chain are -166.16(6)° and 69.1(9)°, respectively. The crystal structure is stabilized by hydrogen bonds along the a and b axes, while the van der Waals forces are the stabilizing interactions in the c direction.

Notes: In a continuation of our program to study the structure-activity relationship of peptide opiates, we report the conformational analysis of two cyclic tetrapeptides related to dermorphin - Tyr-c[D-Orn-Phe-Asp]-NH2 and Tyr-c[D-Asp-Phe-Orn]-NH2. These analogues have similar binding properties marked by a high selectivity for the μ-opioid receptors because of a drastic decrease in the affinity for the δ-opioid receptor. The conformational preferences of these analogues of dermorphin determined from proton nmr, molecular dynamics, and energy minimizations are quite similar. The constraint of the 13-membered ring formed from cyclization is quite evident from the conformational analysis. The constrained ring system acts as a template maintaining the relative orientation of the exocyclic tyrosine and side chain of phenylalanine. Two intramolecular hydrogen bonds measured for the D-Orn analogue in DMSO were disrupted upon the addition of water. For the D-Asp analogue, two intramolecular hydrogen bonds were found stable in DMSO and water. The global conformations of the two peptides determined from nuclear Overhauser effects did not change with the solvent titration. The difference in the hydrogen bonding within the 13-membered ring may account for the slight differences observed in the efficacy of the analogues at the μ-opioid receptors.

Notes: Homonuclear two-dimensional (J,δ) proton spectroscopy has been suggested as a method for the measurement of 1H-31P coupling constants in oligonucleotides. The technique has been applied to a dinucleoside monophosphate G2′p5′C and a deoxydecanucleotide d(ACATCGATGT). PCILO energy calculations have been carried out to find minimum energy conformations with respect to the DNA backbone torsion angle ε, and these have been considered for the interpretation of the observed H3′-31P coupling constants in oligonucleotides.

Notes: Constrained conformational energy minimizations have been used to calculate an adiabatic (φ, ψ) potential energy surface for the disaccharide sucrose. The inclusion of molecular flexibility in the conformational energy analysis of this disaccharide was found to have a significant effect upon the allowed conformational space of the molecule. Three low-energy regions were identified on the adiabatic energy surface, and two of these regions were found to contain two related local minimum-energy conformations, with similar energies, differing only in the directionality of the intra-residue hydrogen bonds of the glucose portion of the molecule. The known crystal structures of seven molecules containing the sucrose moiety all fall within the region of the primary allowed minimum and are consistent with the relaxed energy map, while these crystal conformations could not be rationalized using energy maps for rigid residue geometries. The greater flexibility of the furanoid ring relative to that of the pyranoid ring contributed significantly to the enlargement of the low-energy region on the adiabatic map. However, in spite of the importance of limited flexibility in understanding the conformation and fluctuations of sucrose, this molecule was found to be considerably more rigid that some other disaccharides, such as maltose and cellobiose, in accord with experimental studies.

Notes: Molecular dynamics simulations have been used to study the motions in vacuum of the disaccharide sucrose. Ensembles of trajectories were calculated for each of the five local minimum energy conformations identified in the adiabatic conformational energy mapping of this molecule. The model sucrose molecules were found to exhibit a variety of motions, although the global minimum energy conformation was found to be dynamically stable, and no transitions away from this structure were observed to occur spontaneously. In all but one of these vacuum trajectories, the intramolecular hydrogen bond between residues was maintained, in accord with recent nmr studies of this molecule in aqueous solution. Considerable flexibility of the furanoid ring was found in the trajectories. No “flips” to the opposite puckering for this ring were found in the simulations starting from the global minimum, although such a transition was observed for a trajectory initiated with one of the higher local minimum energy conformations. Overall, the observed structural fluctuations were consistent with the experimental picture of sucrose as a relatively rigid molecule.

Notes: Current ideas on unfolding equilibria in two-chain, coiled-coil proteins are examined by studies of a species of ββ tropomyosin that is sulfhydryl blocked at C190 and disulfide cross-linked at C36 (\documentclass{article}\pagestyle{empty}\begin{document}$ ^. \beta \_\beta ^. $\end{document}). The desired species is produced by a seven-step process: (1) Rabbit skeletal muscle, comprising predominantly αα and ββ species, is oxidized with ferricyanide, cross-linking both species at C190. (2) The product is carbamylated at C36 of β chains, using cyanate in denaturing medium at pH 6. (3) All C190 cross-links are reduced with dithiothreitol (DTT). (4) All C190 sulfhydryls are permanently blocked by carboxyamidomethylation. (5) Chromatography on carboxymethylcellulose in denaturing medium is used to separate C190-blocked α chains from C190-blocked, C36-carbamylated β chains. (6) The latter are decarbamylated in denaturing medium by raising the pH to 8.0. (7) The C190-blocked β chains are renatured and cross-linked at C36 by ferricyanide. The procedure and the quality of the final product are judged by NaDodSO4/polyacrylamide gel electrophoresis, titration of free sulfhydryls, and electrophoretic analysis of trypsin digestion products. Thermal unfolding curves are reported for the resulting pure \documentclass{article}\pagestyle{empty}\begin{document}$ ^. \beta \_\beta ^. $\end{document} species and for its DTT-reduction product. The latter (\documentclass{article}\pagestyle{empty}\begin{document}$ ^. \beta \beta ^. $\end{document}) show equilibrium thermal unfolding curves that are very similar to those of the parent ββ noncross-linked species. The \documentclass{article}\pagestyle{empty}\begin{document}$ ^. \beta \_\beta ^. $\end{document} cross-linked species unfolds in a single-phase, cooperative transition with a melting temperature intermediate between the pretransition and posttransition shown by its cross-linked counterpart, the C190 cross-linked, C36-blocked species (\documentclass{article}\pagestyle{empty}\begin{document}$ .\beta^- \beta .$\end{document}), which was studied earlier. These transitions are compared with one another and with that of the doubly cross-linked species, , in the light of two extant physical models for such transitions. The all-or-none segments model successfully rationalizes the data qualitatively for the \documentclass{article}\pagestyle{empty}\begin{document}$ .\beta^- \beta .$\end{document} and \documentclass{article}\pagestyle{empty}\begin{document}$ ^. \beta \_\beta ^. $\end{document} species if the usual postulates of greater inherent stability of the amino vs the carboxyl end of the molecule and of strain at each cross-link are accepted. However, the same model then requires that the species be the least stable of the three, whereas experiment shows the opposite, thus falsifying the all-or-none segments model. The continuum-of-states model is also qualitatively in accord with data on the \documentclass{article}\pagestyle{empty}\begin{document}$ .\beta^- \beta .$\end{document} and \documentclass{article}\pagestyle{empty}\begin{document}$ ^. \beta \_\beta ^. $\end{document} species. In fact, the general features of the transition in C36 cross-linked vs C190 cross-linked species were predicted by a statistical mechanical theory embodying the continuum-of-states model for singly cross-linked species. Moreover, since the same theory avers that loop entropy greatly stabilizes the large region between cross-links in the species, perhaps offsetting the effect of strain, qualitative considerations alone are insufficient to falsify this model in the face of the data on doubly cross-linked species. Thus, one model can be eliminated, but the second cannot. However, until quantitative simulations are done and found to agree with these data, the continuum-of-states model must still be considered questionable.

Notes: A novel application of dynamic programming to the folding problem for RNA enables one to calculate the full equilibrium partition function for secondary structure and the probabilities of various substructures. In particular, both the partition function and the probabilities of all base pairs are computed by a recursive scheme of polynomial order N3 in the sequence length N. The temperature dependence of the partition function gives information about melting behavior for the secondary structure. The pair binding probabilities, the computation of which depends on the partition function, are visually summarized in a “box matrix” display and this provides a useful tool for examining the full ensemble of probable alternative equilibrium structures. The calculation of this ensemble representation allows a proper application and assessment of the predictive power of the secondary structure method, and yields important information on alternatives and intermediates in addition to local information about base pair opening and slippage.The results are illustrated for representative tRNA, 5S RNA, and self-replicating and self-splicing RNA molecules, and allow a direct comparison with enzymatic structure probes. The effect of changes in the thermodynamic parameters on the equilibrium ensemble provides a further sensitivity check to the predictions.

Notes: The potential of mean force (PMF) approach for treating polyion-diffuse ionic cloud interactions [D. M. Soumpasis (1984) Proceedings of the National Academy of Sciences USA 81, 5116-5120] has been combined with the AMBER force field describing intramolecular interactions. The resultant generalized AMBER-PMF force field enables one to treat the conformational stabilities and structural transitions of charged biomolecules in aqueous electrolytes more realistically. For example, we have used it to calculate the relative stabilities of the B and Z conformations of d(C-G)6, and the B and heteronomous (H) conformations of dA12 · dT12, as a function of salt concentration. In the case of d(C-G)6, the predicted B-ZI transition occurs at 2.4M and is essentially driven by the phosphate-diffuse ionic cloud interactions alone as suggested by the results of earlier PMF calculations. The ZII conformer is less stable than the B form under all conditions. It is found that the helical parameters of the refined B and Z structures change with salt concentration. For example, the helical rise of B-DNA increases about 10% and the twist angle decreases by the same amount above 1M NaCl.In the range of 0.01-0.3M NaCl, the H form of dA12 · dT12 is found to be more stable than the B form and its stability increases with increasing salt concentration. The computed greater relative stability of the H conformation is likely due to noninclusion of the free energy contribution from the spine of hydration, a feature presumed to stabilize the B form of this sequence.

Notes: Poly(O-benzyl-L-serine) of degree. polymerization 90 in oriented film cast from dichloroacetic acid (DCA) solution yielded the x-ray patterns of the intramolecular cross-β form, in which the extended segments stand upward and downward to the surfaces of a thin film. These x-ray photographs further suggest that residual solvent molecules are bound to the polymer, and it may provide an important clue in understanding the nature of the cross-β conformation in a diluted DCA solution.

Notes: Despite its antiparallel symmetry, DNA often appears to possess a permanent electric dipole moment in transient electro-optical experiments. We propose that this may be due to the asymmetric binding of charged ligands to the DNA. We have used the fluctuating dipole theory of Kirkwood and Shumaker to calculate the contribution of asymmetric ligand binding to the electro-optic orientation function, and Monte Carlo computer simulation to calculate the reversing pulse behavior, as a function of ligand binding density. The results indicate that the effect should be observable even against the background of the sizable induced dipole moment produced by polarization of the counterion atmosphere.

Notes: Examination of binding information in the form of derivative (or finite difference) measurements is explored (1) experimentally by a thin-layer optical procedure (Dolman, D. & Gill, S. J. (1978) Anal. Biochem. 87, 127-134) and (2) theoretically by simulation in order to determine the influence of the number of data points and their standard error upon the resolvability of binding parameters in cooperative and non-cooperative systems. The data is described by the difference in optical absorbance divided by the change in the logarithm of the ligand activity and each data point is assumed to be influenced by a random error with a given variance. It is found that increasing the number of data points, which in turn effectively reduces the magnitude of the observed absorbance changes, results in an increase in the uncertainty of the resolved parameters of the system. The effect is verified by both experimental and simulation studies. Thus one is led to suggest that fewer measurements for the change of absorbance with larger magnitudes produces the most favorable situation for parameter resolution when the data is in the form of finite difference measurements.

Notes: The dynamic mechanical properties of elastin have been quantified over a temperature and hydration range appropriate for a biological polymer. Composite curves of the tensile properties at constant water contents between 28.1 and 44.6% (g water/100 g protein) were typical of an amorphous polymer going through its glass transition. Water content had no effect on the shape of the curves, but shifted them a distance aC along the frequency axis. The combined effects of hydration and temperature are given in a series of isoshift curves where elastin's properties are constant along any one curve. A 1% change in hydration has the same effect as a 1°-2° change in temperature, depending on the initial water content and temperature. Theoretical isoshift curves that matched the experimental data were predicted using the WLF equation and coefficients determined from the data. These data form a basis to predict the role of elastin in arterial disease based on changes in its chemical and physical environment.

Notes: A simple theory is developed for the extension of a uniformly charged linear polyion attached at one end in an electric field. For a polyion consisting of a very large number (N) of Kuhn lengths (b), the mean extension in the direction of the electric field (E) is given accurately by Rz = (b/A) In (sinh (NA)/NA), where A = EQb/kBT and Q is the effective charge associated with each Kuhn length. For any value of E, no matter how small, a polyion of sufficient length, such that NA ≫ 1.0, will be essentially fully extended. When A ≪ 1.0, as is the case for DNA in weak electric fields, the head end of the chain is only weakly oriented, even though the stem and tail may be completely aligned. In the linear regime, NA 〈 1.0, Rz is proportional to E, Q, and N2. The linear density of chain elements is calculated for a chain in the linear regime. This theory provides a basis to determine the product Qb from direct microscopic measurements of Rz vs. E for fluorescent-labeled circular DNAs that are topologically snared near voids or surfaces of gels. Experimental realization of this suggestion is described in the following paper.

Notes: Individual ethidium-stained DNA molecules, embedded in an agarose gel made with electrophoresis buffer (0.05 molar salt), are observed using a fluorescence microscope. In the first experiment, open circular 66 kilobase pair (kbp) plasmids, immobilized by agarose fibers threaded through their centers, display entropic “rubber” elasticity. The charged molecules extend in an electric field of several volts per centimeter and contract to a compact random coil when the field is removed. The extension of the plasmids as a function of field strength is consistent with the freely jointed chain model when the effective electrophoretic charge density is set at 15 e- per persistence length. In a second experiment, stained linear 48.5 kbp DNA molecules are observed as random coils immobilized in agarose. A measure of their size, here named the “maximal-X-extent,” is taken for 100 molecules and found to average 1.47 μ. A Monte Carlo computer simulation of random coils (freely jointed chain model) gives the same maximal-X-extent value when the persistence length is set at 0.08 μ.

Notes: The temperature dependence of hydrogen isotope exchange rates for lysozyme in 5 molal aqueous glycerol and for poly (D,L-alanine) in a range of glycerol concentrations from 0 molal to 8 molal have been determined. The activation enthalpy of base-catalyzed exchange for poly (D,L-alanine) in water is 4 kcal/mol and passes through a minimum at about 2 molal glycerol before returning to a value of 4 kcal/mol at 4 molal glycerol. Exchange rates for lysozyme have been analyzed with transition state and Kramers's theories. The activation parameters for exchange of protons in lysozyme in the presence of 5 molal glycerol show a similar qualitative behavior to those determined for exchange in the absence of glycerol [R. B. Gregory et al. (1982) Biochemistry 24, 6523-6530]. The activation enthalpies and entropies for the fast-exchanging protons show a gentle increase as H(t), the number of hydrogens remaining unexchanged, decreases. By contrast, the activation parameters for the slowest exchanging protons [H(t) 〈 20] increase dramatically as H(t) decreases. As in water, the activation parameters for exchange of the fast- and slow-exchanging protons in glycerol solution are characterized by two distinct compensation temperatures (510 ± 100 K for the fast protons and 340 ± 40K for the slow protons). These values are not significantly different from those determined for exchange in water.The activation parameters, ΔH

Notes: Two dielectric relaxation peaks were found in moist collagen by the time domain reflectometry. The low-frequency peak around 100 MHz moves little as the water content is varied. Its relaxation strength depends on the content and vanishes for completely dried collagen. This process is concluded to be due to water molecules strongly bound to the tropocollagen. Amount of the bound water is estimated as 0.12 g water/g collagen. Twenty-one water molecules are bound to one repeat of the triple helix. The existence of stringlike water chains is suggested. If the water content is less than 0.5 g water/g collagen, the high frequency peak locates between those of bound and bulk water. Water among the tropocollagen is weakly bound to the collagen. In the higher region it does not change much with the content, being close to that of bulk water. The bulk water appears in this region.

Notes: We have examined the free energy effects of 5-methylation of cytosine on the B ↔ Z conformational equilibrium in DNA. Free energy differences were calculated using the free energy perturbation approach, which uses an easily derived equation from classical statistical mechanics to relate the free energy difference between two states to the ensemble average of the potential energy difference between the states. Calculations were carried both in explicit solvent and (for comparison) in vacuo. The free energy values obtained for the explicit solvent systems are total free energies, with contributions from all parts of the system (solvent + solute), and so are relevant to the B ↔ Z transitions observed under real(physiological) conditions. We calculate that in solution, methylation makes the B → Z transition more favorable by about -0.4 kcal/mole base pair (bp) in free energy. This value compares well with approximate experimentally derived values of about -0.3 kcal/ mole-bp. We also discuss a method for determining the free energy difference between conformational states poorly maintained by a potential energy model. Finally, the effects of methylation on the melting temperature of DNA are examined.

Notes: Molecular dynamics simulations of methyl-β-D-glucoside in water are performed. Although the tg conformation of the hydroxymethylgroup is found to be the lowest in energy for the isolated molecule, it appears to be unstable in water. The well-known absence of a significant amount of the tg conformation in polar solvents can therefore be explained as a solvent effect.

Notes: The coagulation mechanism of thrombin- and reptilase-activated fibrinogen is the focus of this report. Three types of gels were evaluated for their sensitivity to physiologic levels of Ca(II) and Zn(II): Fibrin gel formed directly by adding thrombin or reptilase to fibrinogen, protofibrin gel generated by adding Ca(II) and Zn(II) to soluble fibrin oligomers (protofibrills), and gels induced by sudden pH increase of fibrin monomer (pH 4.9-7.4). The turbidity and viscoelasticity of reptilase- and thrombin-activated fibrin are similarly sensitive to Ca(II) and Zn(II) scanning electron micrographs (SEM) of fibrin gels from pH jump experiments show that their constituent fibers are relatively homogeneous and only mildly affected by divalent cations. This contrasts with the cation sensitivity of normally formed fibrin and protofibrin gels. Based on these and previously published experiments, a graphic shorthand is developed to describe the directionality of linear and lateral polymerization modes that result in cation-driven (proto)fibrin gelation.

Notes: A theory is developed for dynamic light scattering (DLS) from rigid double spirals by treating an invisible rigid cylinder with two helical scattering stripes on opposite sides of its cylindrical surface. The exact initial, or first cumulant, diffusion coefficient Dapp(K) is obtained in terms of the translational diffusion coefficients (D∥ and D⊥) parallel and perpendicular to the symmetry axis, the rotational diffusion coefficients (DR∥ and DR⊥) around the symmetry and transverse axes, the length (L) and radius (b) of the cylindrical surface bearing the stripes, and the pitch (p). Interference effects, namely geometrical antiresonances, between strands, produce deep minima in the static structure factor S(K) and corresponding prominent peaks in Dapp(K). These peaks in Dapp(K) depend sensitively on the rotational dynamics around the symmetry axis, and nearly vanish when DR∥ = 0. Some results for single spirals are also presented. A simpler model in which scattering points are attached at opposite ends of an otherwise invisible thin rigid rod is also treated, and shown to exhibit modest minima in S(K) and corresponding maxima in Dapp(K). Confining this rod to a plane containing K enhances the amplitudes of the oscillations in S(K) and Dapp(K), as expected.Rigid double spirals are employed as crude models for interwound supercoiled DNAs in order to assess the possible occurrence of interference effects. Although native supercoiled DNAs exhibit a cylinder diameter that is much too small to exhibit geometrical antiresonances in the presently accessible range of K2, nearly relaxed supercoiled DNAs are predicted to exhibit their first maximum in Dapp(K) just inside this range. Previously reported data for the effect of Escherichia coli single-strand binding (ssb) protein on the DLS of supercoiled pBR322 DNA cannot be mimicked by a gradual homogeneous reduction of superhelix density with increasing ssb, but instead can be mimicked by inhomogeneous all-or-none binding in which uncomplexed native DNAs and nearly relaxed saturated ssb/DNA complexes coexist in varying proportions. Experimental Dapp(K) and S(K) data for a sample of relaxed pUC8 dimers display, respectively, a broad maximum and a corresponding minimum, in qualitative agreement with rough theoretical predictions.

Notes: Fluorescence of (+)-anti-benzo (a) pyrene diol epoxide [(+)-anti-BPDE] covalently bound to poly (dG-dC) has been studied with steady-state and time-resolved techniques. Extensive formation of excimers is found, even at small (0.008) BPDE/nucleotide ratios. This indicates favored covalent binding to bases close to already modified guanines. Both fluorescence excitation spectra and lifetime measurements reveal two populations of (+)-anti-BPDE adducts: one that can form excimers and one that cannot. Three excimer lifetimes (4.5, 29, and 83 ns) are observed. Differently shifted monomer and excimer excitation spectra are discussed in terms of pyrene-pyrene exciton interactions, consistent with a distance shorter than 7 Å between the excimer-forming BPDE chromophores.

Notes: Being interested in possible effects of sequence-dependent hydration of B-DNA with mixed sequence in fibers, we performed a series of Monte Carlo calculations of hydration of polydeoxyribonucleotides in B form, considering all sequences with dinucleotide repeat. The computational results allow the ten base-stacking types to be classified in accordance with their primary hydration in the minor groove. As a rule, the minor groove is occupied by two water molecules per base pair in the depth of the groove, which are located nearly midway between the planes of successive base pairs and symmetrically according to the dyad there. The primary hydration of the major groove depends on the type of the given base pair. The coordinates of 3 water molecules per base pair in the depth of the major groove are determined by the type of this pair together with its position and orientation in the helix, and are practically independent on the adjacent base pairs. A/T-homopolymer tracts do not fit into this hydration pattern; the base pair edges are hydrated autonomously in both grooves.Analysis of the Li-B-DNA x-ray diffraction intensities reveals those two water positions in the minor groove. In the major groove, no electronic density peaks in suffiecient distance from the base edges were found, thus confirming the absence of any helical invariance of primary hydration in this region.With the help of the rules proposed in this paper it is possible to position the water molecules of the first hydration shell in the grooves of canonical B-DNA for any given sequence.

Notes: A general method is presented for constructing a potential function for approximate conformational calculations on globular proteins. The method involves solving a nonlinear program that seeks to adjust the potential's parameters in such a way that a minimum near the native remains a minimum and does not move far away, while any alternative minima shift so as to remain local minima but eventually rise higher than the level of the near-native minimum. Although the potential trades computational speed for detail by representing each amino acid residue as only a single point, correct secondary structural preferences and reasonable tertiary folding can be built into the potential in an entirely routine way. The potential has been parameterized to agree with the crystal structure of avian pancreatic polypeptide (having 36 residues) in the sense that the lowest minimum found (-407 arbitrary units) is reasonably close to the native (1.8 Å rms interresidue distance deviation). In contrast, the lowest nonnative conformation found after extensive searches by a variety of methods was -399 units and 7.5 Å away. Such potentials may prove to be useful in predicting approximate tertiary structure from amino acid sequence, if they can be generalized to apply to more than one protein.

Notes: Deuterium exchange of 8C protons of adenine and guanine in nucleic acids is conveniently monitored by laser Raman spectrophotometry, and the average exchange rate so determined (〈kA + kG〉) can be exploited as a dynamic probe of the secondary structure of DNA or RNA [J. M. Benevides and G. J. Thomas, Jr. (1985) Biopolymers 24, 667-682]. The present work describes a rapid Raman procedure, based upon optical multichannel analysis, which permits discrimination of the different 8CH exchange rates, kA of adenine and kG of guanine, in a single experimental protocol. For this procedure, simultaneous measurements are made of the intensity decay or frequency shift in separately resolved Raman bands of adenine and guanine, each of which is sensitive only to 8C deuteration of its respective purine. Resolution of the rates kA and kG is demonstrated for the mononucleotide mixtures, 5′-rAMP + 5′-rGMP and 5′-dAMP + 5′-dGMP, for the polynucleotides poly (dA-dT) . poly (dA-dT) and poly (dG-dC)·poly (dG-dC), for calf thymus DNA, and for the 17 base-pair operator OR3. We show that the different exchange rates of adenine and guanine, in nucleotide mixtures and in DNA, may also be calculated independently from intensity decay of the composite 1481-cm-1 band, comprising overlapped adenine and guanine components, over a time domain that encompasses two distinct regimes: (1) a relatively more rapid exchange of guanine, and (2) a concurrent slower exchange of adenine. Both methods developed here yield consistent results. We find, first, that exchange of guanine is approximately twofold more rapid than that of adenine when both purines are present in the same structure and solvent environment, presumably a consequence of the greater basicity of the 7N site of guanine. Second, we find that adenine suffers greater retardation of exchange than guanine when both purines are incorporated into a “classical” B-DNA secondary structure, such as that of calf thymus DNA. This finding suggests different microenvironments at the 7N-8C loci of adenine and guanine in aqueous B-DNA. We also confirm that adenine residues of B-form poly (dA-dT)·poly (dA-dT) exchange much more slowly than those of other B-DNA sequences, implying a secondary structure for the alternating-AT sequence with unusual stereochemistry in the major groove. The greater resistance of adenine than guanine to 8CH exchange in the B-DNA secondary structure is more evident in high molecular weight calf thymus DNA and in the alternating AT and GC copolymer duplexes than in the smaller 17 base-pair operator OR3. The present results provide direct experimental evidence for structural heterogeneity of B-DNA in aqueous solution, involving the purine 7N-8C imidazole network. The methods developed here establish an experimental basis for future study of purine-specific interactions that may be important in gene regulatory complexes and in nucleic acid condensation and packaging processes.

Notes: A new block-units method for rapid conformational calculation of large nucleic acid fragments has been developed. Atomic structure of polynucleotides has been approximated by the block-units structure. Each monomer of the polynucleotide consists of three one-center block units: phosphate, ribose, and nucleic base. Full conformational energy of the polynucleotide is separated into two parts. The first part is the energy of the short-range interactions between adjacent block units and is calculated on the basis of the potential energy surface model of the dinucleotide fragment pXp (where × = A, G, T, U, C). The second part is the energy of the middle- and long-range interactions between separated block units, and is calculated as a sum of the effective interaction energies between centers of the block units. The present block-units method is in agreement within the range of ±0.5 kcal/mol with the method of the atom-atom potentials, but the former is 30-100-fold faster. The block-units method is recommended for screening of the probable conformations of the large polynucleotide systems.

Notes: The two-level hierarchical methodology is suggested for conformational calculations of large fragments of nucleic acids. The method of the first level is intended for performing a fast screening of the conformational phase space. The high-level method may be used to refine structurally important conformations. The method of the first level is the block-units method, which has been developed specially for these purposes (see part I). It has been shown that the block-units method allows the satisfactory calculation of the structure parameters of the optimal conformations of polynucleotides. The results of the conformational rerrangement calculations of the TψC loop of the tRNAPhe after modification of its sequence are represented.

Notes: Analogous with the Potts model that describes the helix-coil transition in the isolated polypeptide chain (a Hamiltonian model allowing for the energy U of hydrogen bond formation) the number Q of conformational states of a repeating unit of the chain and the topology of Δ = 3 hydrogen bond formation (the hydrogen bond fixing three pairs of ϕψ chain rotations) has been constructed and the corresponding transfer-matrix has been obtained. In the thermodynamical limit, the partition function is expressed through the principal root of the cubic equation. The degree of helicity, the transition point and range, the correlation length, the number of junctions between the helical and coiling sections as well as the mean length of helical and coiling sections are calculated. Empirically introduced parameters of the Zimm-Bragg theory, constants of hydrogen bond formation s, and the cooperativity parameter σ as functions of microscopic parameters U, Q, and Δ are obtained by direct calculations. The behavior of this model was investigated at other topologies of the hydrogen-bond closing Δ = 2 and Δ = 4, and it was suggested that the actual polypeptide chain (Δ = 3) provides the optimum correlation of helical structure of the order of globule dimensions. An expression was obtained for the maximum correlation length of the order ξ ∼ Q(Δ-1)/2. For a System with solvent competing for the formation of hydrogen bonds with peptide groups a Hamiltonian model was constructed that took account of the energy E of the formation of hydrogen bond with the solvent and the number q of orientations of a solvent molecule about the peptide groups. It is shown that by the redefinition of the temperature parameter, the model with solvent reduces to the model of an isolated chain. Aside from the definition relationship that exists between the parameters of the theory U 〈 2E 〈 Uq and the ordinary helix-coil transitions (“melting”), the model also describes the transition from the coiling state to the helical one (“arrangement”) under heating. The change in temperature and transition range with solvent parameters was discussed and it was shown, that despite the difference in ΔT for the given polypeptide chain (Q = constant) with different solvent parameters, at “melting” and “arrangement,” the transition occurred at the same correlation length (the same cooperativity).

Notes: This paper reports the chiroptical properties of thionated N-acyl amino acid and N-acyl dipeptide N′-methylamide models. It was found that the optical activity of the thioamide chromophore is dominated by the chiral contribution of perturbants attached to Cα at the N-H side of the thioamide group. The appearance of a strong negative ππ* band near 270 nm is indicative of the semiextended conformation of this residue. The φ ∼ -70°, ψ ≥ 120° set of torsion angles is compatible with a type II βt-turn or a γt-turn conformation with the perturbing N-H side residue in the i + 1 position of the turn. (The subscript t or tt denotes that one or both of the H-bonded moieties is thioamide.) Earlier data show that both βt- and γt-turns may be fixed by C=S⃛H-N(CO) intramolecular H bonds. The appearance of one or two weak nπ* bands and a positive ππ* band at about 270 nm is characteristic of type II βt-turns containing the H-bonded thioamide group attached to the glycine residue in position i + 2. The extended conformation (φ ∼ -140°, ψ ∼ 140°) of a residue after the thioamide group gives rise to a negative nπ* and a positive ππ* band of comparable magnitude. Peptid1e sequences with alternating thioamide-amide-thioamide backbone tend to adopt 1t ⇆ 4t H-bonded βtt conformations. CD studies show that type II βtt-turns have unique chiroptical properties: the ππ* region is dominated by an exceedingly strong negative band near 260 nm (|Δε| = 19-24) accompanied by a weaker band at higher wavelength values.

Notes: Molecular dynamics simulations on the transmembrane antibiotic peptide alamethicin have been performed in the NVT ensamble (i.e., the number of particles N, the volume V, and the temperature T of the system are kept constant). Results on the structure and conformational flexibility of this molecule are discussed and compared with previous experimental CD, x-ray, nmr data and theoretical computations on fragments analogues.An extensive study of structural and dynamic properties from H-bonding pattern analysis is presented. Evidences for a largely α-helix structure with some extent of freedom in the C-terminal region are found.Further, a partition of the molecule into three regions on the base of structural features and dynamic behavior has been proposed, and the correlation among the motions of the three regions is described.

Notes: The effect of substituents (X = H, Me, or F at C(6), R = H or Me at C(2′) of the allyl side chain) on the photoisomerization (λ = 350 nm) of 6-allylcyclohex-2-enones 1 in MeCN is studied. Substituents X control the overall efficiency of intramolecular [2 + 2] photocycloadduct formation (Φ: Me 〉 F 〉 H) but do not exercise an influence on the orientation of addition of the exocyclic double bond to the enone C=C bond. In contrast, replacement of the prop-2-enyl (R = H) by a 2-methylprop-2-enyl (R = Me) side chain causes a change in the tricyclo[3.3.1.02,7]nonan-6-one 4 vs. tricyclo[4.2,1.03,8]nonan-7-one (5) product ratio from 100:0 (R = H) to roughly 2:1 (R = Me) but has almost no bearing on the relative rates of conversion of 1 to products. For C(6)-unsubstituted enones 1aa and 1ba (X = H), the efficiency of cyclization becomes low enough so that lumiketone rearrangement to bicyclohexanones 6 and 3-isopropylcyclopent-2-enones 9 becomes competitive. Enones 9 undergo consecutive intramolecular [2 + 2] photocycloaddition to tricyclo[3.2.1.03,6]octan-2-ones 7 and to tricyclo[3.2.1.03,6]octan-7-ones 8, compounds 8 only being formed when R = Me.

Notes: Substituent, Heteroatom, and Solvent Effects on the Thermal-Bleaching Kinetics and Absorption Spectra of Photomerocyanines Issued from Spiro[indoline-oxazines]Quantitative information useful for the development of new photochromic systems is obtained from the study of heteroatom and substituent effects on the thermal-bleaching kinetics and the absorption spectra of the photomerocyanines issued from spiro[indoline-oxazines]. The effect on photochromic properties of the presence of N-atoms either in the dimethine bridge or in the aromatic rings has been investigated through the comparison of spiro[indoline-naphthopyrans] C with spiro[indoline-naphthoxazines] A and with spiro[indoline-quinolinoxazines] B. Besides the occurrence of biexponential thermal-bleaching kinetics in non-polar solvents is observed: a tentative explanation for this observation is given which involves the formation, in either sequential or parallel steps, of energetically distinct stereoisomers of the opened form.

Notes: The kinetics of the electron transfer between reduced spinach [2Fe-2S]-ferredoxin and the optically active complexes [Co((R,R)- or (S,S)-alamp)py]+ (I), [Co((R,R)- or (S,S)-promp)H2O]+ (IIa), and [Co((R,R)- or (S,S)-promp)py]+ (IIb) have been investigated. The reactions are stereoselective, and for I and IIa, the Stereoselectivity strongly depends on temperature due to large differences in the activation enthalpy between enantiomeric reagents. Isokinetic behaviour is observed between enantiomers, the ΔΔHΔ-Λ# values being largely compensated by the ΔΔSΔ-Λ# values. The compensation behaviour is explained by the combination of stereochemical interactions and desolvation processes on formation of the precursor complex or the transition state.

Notes: A stereocontrolled synthetic route to optically pure (-)-(S)-ipsenol (1), the pheromone of Pityokteines curvidens and various other bark-beetle species is described. Key step of the synthesis is an enantioselective aldol reaction using a chiral titanium-carbohydrate complex (Scheme 1). The carboxylate function of the optically pure β-hydroxy acid 5 thus obtained in mol quantities is then elaborated to the diene moiety by standard methodology (Scheme 2).

Notes: Synthesis of 2-Oxo-2H-pyran-5-carboxylate Derivatives3-Substituted diethyl pent-2-enedioates are easily formylated by means of ethyl formate/TiCl4/4-methylmorpholine to produce the ethoxymethylene derivatives, which are smoothly cyclized either with HCOOH or PPA to corresponding 2-oxo-2H-pyran-5-carboxylate derivatives.

Notes: A series of 5-substituted (-)-(S)-N-[(1-ethylpyrrolidin-2-yl)methyl]-2,3-diniethoxybenzanndes were made by reaction of the corresponding benzoyl chlorides with (S)-1-ethylpyrrolidine-2-methylaruine (→ 14-16, 18-21). The acids required were prepared in a regiospecific manner from 5-bromo-2,3-dimethoxybenzoic acid which was protected as dihydrooxazole (→ 4-8), metalated, reacted with various electrophiles (MeI, EtI, BuBr, CC13CCl3 or MeSSMe), and hydrolyzed (→9-13). Alternatively, (-)-(S)-5-bromo-N-[(1-ethylpyrrolidin-2-yl)methyl]-2,3-di-methoxybenzamide was treated with KH followed by BuLi and an electrophile (I2 or Me3SiCl) to give the 5-iodo and 5-(trimethylsilyl) derivatives 17 and 22, respectively. All 5-substituted amides were highly potent inhibitors of [3H]spiperone binding in rat striatal membranes with IC50 values of 0.5 to 5 nM (Table 3). Thus, a relatively large steric bulk can be accomodated in the position para to the 2-MeO group. This work also supports the notion that a positive as well as negative electrostatic potential can be located in this position. A selected number of derivatives were also investigated in vivo and found to inhibit apomorphine-induced behavioural responses in the same dose range as haloperidol and raclopride (Table 4). This new group of benzamides is suitable for investigations of dopamine D-2 receptors in labelled or unlabelled form.

Notes: Chiral ligands coordinated to metal ions exert a selectivity towards the additional coordination of racemic substrates. Experimentally determined equilibria distributions of [Co(L3)2]3+ and [Co(L3)(L2)(X)]n+ are compared with calculated data based on strain-energy minimization (L3: trap = propane-1,2,3-triamine; 1,2,4-trab = butane-1,2,4-triamine; 1,2,3-trab = butane-1,2,3-triamine; 1,3,4-trpe = pentane-1,3,4-triamine; 1,3,4-tmeb = 2-methylbutane-1,3,4-triamine; 1,2,4-trpe = pentane-1,2,4-triamine; L2: en = ethane-1,2-diamine; pn = propane-1,2-diamine; X: NH3, OH2, OH-). Equilibration of Co(III) complexes was achieved by oxygenation of aqueous solutions of Co(II) salts in presence of the ligands. Quantitative isomer distribution was investigated with HPLC, and quantitative analysis of the enantiomeric excess (ee) of the racemic substrate (present in a two-fold excess) was studied, after chromatographical recovery, by 1H-NMR analysis of its Mosher-acid derivative. There is good agreement between calculated and experimental data. Systems with L = 1,2,4-trab are, as expected, relatively poorly discriminating (ee([Co(1,2,4-trab)2]3+) ∼ 5%; ee([Co(1,2,4-trab)(pn)(X)]n+) ∼ 10%). Calculations indicate that Me substitution of the ligand backbone of 1,2,4-trab (and trap) leads to an increased enantioselectivity (with practically constant isomer selectivity), and at the optimum site for substitution ∼ 90% ee is predicted.

Notes: The reaction of 3-(dimethylamino)-2H-azirines 1a-c and 2-amino-4,6-dinitrophenol (picramic acid, 2) in MeCN at 0° to room temperature leads to a mixture of the corresponding 1,2,3,4-tetrahydroquinazoline-2-one 5, 3-(dimethylamino)-1,2-dihydroquinazoline 6, 2-(1-aminoalkyl)-1,3-benzoxazole 7, and N-[2-(dimethylamino)phenyl]-α-aminocarboxamide 8 (Scheme 3). Under the same conditions, 3-(N-methyl-N-phenyl-amino)-2H-azirines 1d and 1e react with 2 to give exclusively the 1,3-benzoxazole derivative 7. The structure of the products has been established by X-ray crystallography. Two different reaction mechanisms for the formation of 7 are discussed in Scheme 6. Treatment of 7 with phenyl isocyanate, 4-nitrobenzoyl chloride, tosyl chloride, and HCl leads to a derivatization of the NH2-group of 7 (Scheme 4). With NaOH or NaOMe as well as with morpholine, 7 is transformed into quinazoline derivatives 5, 14, and 15, respectively, via ring expansion (Scheme 5). In case of the reaction with morpholine, a second product 16, corresponding to structure 8, is isolated. With these results, the reaction of 1 and 2 is interpreted as the primary formation of 7, which, under the reaction conditions, reacts with Me2NH to yield the secondary products 5, 6, and 8 (Scheme 7).

Notes: The intramolecular cycloaddition of the previously described azidoalkene 16, the related diacetates 7 and 13, and the monoacetate 8 led diastereoselectivity to the (2R)- and (2S)-configurated hydropyridotriazoles 17, 9 and 11, 14 and 15, and 10 and 12, respectively (Scheme 1). Thermolysis of 16 gave also the aziridine 18, its proportion increasing with reaction time. The diastereoselectivity of the cycloaddition- is rationalized on the basis of steric interactions and of H—bonds in the transition state. Photolysis in benzene partially transformed 9 into the aziridine 19. Treatment of 9 with aqueous AcOH gave 19 and the tetrahydrofuran 20, with AcOH in benzene 20 and the triacetate 23, and with aqueous H2SO4 in THF, the primary alcohol 22 (room temperature) or 19 and 22 (0°). Deacetylation of 9 followed by reaction with pyridinium hydrochloride led to the tetrahydrofuran 21 and the chloride 24 (Scheme 2). The diacetate 22 and the triacetate 23 gave the tripl 25 which was deprotected to 26. Reduction of the keto-aziridine 18 (NaBH4) gave the alcohols 27 and 29 which were acetylated to give 28 and 19, respectively (Scheme 3). Treatment of the aziridine 28 with AcOH in benzene followed by deacetylation gave 30 and hence 31. AcOH in benzene transformed the triazoline 15 first into the aziridine 32 and hence into 33, which was deprotected to give the triol 34 and hence 35. The 2-(hydroxymethyl)piperidines 26, 31, and 35 inhibited Vibrio cholerae sialidase with K1 = 3.8 · 10-2 M, 3.4 · 10-3 M, and 1.5 · 10-4 M, respectively. The conformation of the glycerol side chain of these compounds and of the unbranched piperidines 2-4 deviates from the one of Neu5Ac (and Neu2en5Ac). This finding is rationalized by an H-bond between OH—C(8) and NH—C(6). The conformations and the K1 values of 26, 31, and 35 correlate with each other.

Notes: 1H-Cyclopropa[g]quinoline (3-aza-lH-cyclopropa[b]naphthalene; 17) was synthesized via interception of the heterocyclic ortho -quinodimethane 15 with l-bromo-2-chlorocyclopropene, followed by aromatization of the adduct 16 with t-BuOK.

Notes: Reaction of the carbanions derived from chloromethyl phenyl sulfone or 1 -chloroethyl phenyl sulfone with the cationic [Fe(arene)Cp] complexes 8 or 9 produced isolable c-adducts 10-12. Attempted base induced elimination of the s̰-adducts, which would have led to products of vicarious nucleophilic substitution (VNS reaction), failed. Similarly, no VNS products were obtained, when the (arene)tricarbonylchromium complexes 4 were reacted with the anion of chloromethyl phenyl sulfone.

Notes: Thiamine hydrochloride (1a; 3-[(4-amino-2-methylpyrimidin-5-yl)methyl]-5-(2-hydroxyethyl)-4-methylthia-zolium chloride hydrochloride; vitamin B1) has been synthesized in excellent yield by condensation of 3-mercapto-4-oxopentyl acetate (5a) with 3, 4-dihydro-7-methylpyrimido[4, 5-d]pyrimidine (4) in formic acid. The two intermediates 5a and 4 are prepared from 3-chloro-4-oxopentyl acetate (3) and 4-amino-2-methyl-5-(aminomethyl)-pyrimidine (Grewe diamine; 2a), respectively.

Notes: Ring Enlargements and Ring Contractions in the Reaction of 1, 3-Oxazolidine-2, 4-diones and l, 3-Thiazolidine-2, 4-dione with 3-Amino-2H-azirinesThe reaction of 3-amino-2H-azirines 1 and 1, 3-oxazolidine-2, 4-diones 2 in MeCN at room temperature leads to 3, 4-dihydro-3-(2-hydroxyacetyl)-2H-imidazol-2-ones 3 in good yield (Scheme 2, Table 1). A reaction mechanism proceeding via ring enlargement of the bicyclic zwitterion A to give B, followed by transannular ring contraction to C, is proposed for the formation of 3. This mechanism is in accordance with the result of the reaction of 2a and the 15N-labelled 1a*: in the isolated product 3a*, only N(3) is labelled (Scheme 1). The analogous reaction of 1 and 1, 3-thiazolidine-2, 4-dione (5) is more complex (Schemes 4 and 5, Table 2). Besides the expected 3, 4-dihydro-3-(2-mercaptoacetyl)-2H-imidazol-2-ones 7, 5-amino-3, 4-dihydro-2H-imidazol-2-ones of type 8 and/or N-(1, 4-thiazin-2-ylidene)ureas 9 are formed. In the case of 2-(dimethylamino)-1-azaspiro[2. 3]hex-1-ene (1d), the postulated eight-membered intermediate 6d could be isolated. Its structure as well as that of 9f has been determined by X-ray structure analysis. A reaction mechanism for the formation of the 1, 4-thiazine derivatives of type 9 is proposed in Scheme 6.

Notes: The synthesis of the D-xylopyranose-5-spiro-1′-cyclopropane 5, its methyl α-D-glycoside 7 and its benzyl β-D-glycoside 13 from D-glucose is described, and their conformation in solution is discussed. A Königs-Knorr glycosidation of 10 reveals the ionic intermediate of a 1, 1-(dibromocyclopropyl)carboxonium ion type to be stable against opening of the cyclopropane ring. Very weak inhibition of saccharase was observed for the α-D-configurated methyl glycoside 7, whereas the β-D-configurated benzyl glycoside 13 did not inhibit emulsin.

Notes: No Abstract.The English Footnotes (*) referring to Schemes 1-6 are intended to provide an extension of this summary. In the Footnote (*) to Scheme 5, a definition of the term ‘chirogenic reaction step’ is given.

Notes: Electrophilic Substitution Reactions of 1,1-;Difluoro-1H-cyclopropabenzene1,1-Difluoro-1H-cyclopropabenzene (1) can be deprotonated with strong bases at C(2). The resulting 1,1-di-fluoro-2-lithio-1H-cyclopropabenzene (2) reacts with electrophiles to form C(2)-substituted derivatives of 1. The Diels-Alder reactions with electron-poor dienes, characteristic for 1H-cyclopropabenzene, do not occur with the 1,1-difluoro analogue 1.

Notes: A synthesis of Methylated Epoxyhydroazulenones by Intramolecular [4 + 3] Cycloaddition of an Oxyallyl Intermediate, Generated from 1,1-Dichloro-6-(3-methyl-2-furyl)hexan-2-one(Z)-3-Methylpent-2-en-4-yn-1-ol (7) was transformed to 2-(4-chlorobutyl)-3-methylfuran (4b) and 2-(but-3-enyl)-3-methylfuran (10a) by C-alkylation and 5-exo-dig cyclization. The Grignard derivative formed from 4b gave 1,1-dichloro-6-(3-methylfur-2-yl)hexan-2-onc (1b) on reaction with dichloroacetyl chloride. This dichloromethyl ketone undergoes a base-induced cyclization to form diastereoisomeric 7-chloro-1,2,3,6,7,8a-hexahydro-4-methyl-8H-3a,6-epoxyazulen-8-ones (3bα and 3bβ) by way of an intramolecular [4+3] cycloaddition of an oxyallyl intermediate 2b. By dechlorination and hydrogenation of 3bβ, the tricyclic hydroepoxyazulenones 18 and 19 have been synthesized.

Notes: The marine sponge Spongia zimocca SCHMIDT, 1862, collected in front of the torrent II Rogiolo, south of Livorno, contains the sesquiterpene rogiolol acetate (= (+)-(2R,3S,6R,8R,9R)-2,8-dibromo-9-chloro-1,1,9-trimethyl-5-methylidenespiro[5.5]undec-3-yl acetate; (+)-3a), which represents the first chamigrane isolated from a sponge. Although compounds of this class are common in red seaweeds of the genus Laurencia, and our sponge actually contains 9-bromochamigrene and a variety of other metabolites of nearby growing Laurencia sp., (+)-3a is unique to our sponge.

Notes: The stereochemical outcome of the base-catalyzed cyclization of diketones 5-8 has been investigated under protic conditions (Scheme 3). The more stable trans-fused ketols are preferentially formed in kinetically controlled aldol reactions, when the incipient angular substituent R = H (6 → 10a) or CN (7 → 11a, 8a → 12a). For R = Me (as in 5), axial attack of the side-chain enolate double bond on the ring C≡O group results in the rather selective formation of cis-9b. It is assumed that these cyclizations are controlled by relative product stabilities (product-like transition state) and steric effects. The competition between fused (e.g. 9) and bridged ketol (e.g. 13) formation in these cyclizations is discussed. The cis-fused (‘steroid’) ketols were readily equilibrated with their trans-counterparts (9b ⇄ 9a, 10b ⇄ 10a, 11b ⇄ 11a) under aprotic conditions (5 mol-% of LDA, THF, 0°), thus, allowing assessments of relative stabilities.

Notes: The photo-oxygenation of adamantylideneadamantane (1) on siliceous supports using admixed granules of ion-exchange resin fixed to methylene blue (MB) and rose bengal (RB) gave exclusively the corresponding dioxetane derivative 2 for the former sensitizer, while the latter gave 2 and traces of the epoxide 3. RB and the charge-transfer complex produced from N-ethylcarbazole and 2,4,5,6-tetranitrofluoren-9-one both reacted with chemically generated singlet oxygen to give superoxide radical anion. Trapping of the latter with 5,5-dimethyl-1-pyrroline 1-oxide gave an adduct exhibiting a characteristic ESR spectrum. The treatment of 1 in MeOH with 30% aqueous H2O2 for 22 h at 60° gave 3 in 100% yield. Repetition of this experiment in the presence of 2,6-di(tert-butyl)-p-cresol caused no significant change. These results indicate that singlet oxygen reacts with 1, in the presence of RB, by two different processes. The first leads to dioxetane formation. The second process involves conversion of singlet oxygen by RB to superoxide radical anion which subsequently gives H2O2 so producing epoxide 3 from 1.

Notes: Reduction of 6 by borocyanohydride yielded the new dihydroflavin 7. The intermediate product 8 could only be observed in solution by 1H-NMR. The chemical and physical properties of 7 are reported. UV/VIS, 1H- and 13C-NMR, and luminescence techniques were used.

Notes: Benzobenzvalene (naphthvalene; 1) is shown to add SO2 to a lateral bicyclobutane bond with formation of a sulfone 2 and a ‘γ-sultine’ 3. The structure of the latter is unambiguously established by X-ray diffraction. Both adducts extrude SO2 upon direct photolysis at 254 nm and regenerate 1 accompanied by naphthalene in a 1:3 ratio. This result is interpreted in terms of a reversible homolytic cleavage leading, for both, 2 and 3, to the same sulfinyloxy biradical 5, which by loss of SO2 gives the benzoprefulvene biradical 6. The latter in its singlet state undergoes ring closure to 1, or it opens to give naphthalene.

Notes: Phenol, 4-methoxyphenol, 4-nitrophenol, methyl orsellinate (1), and 2,6-di(tert-butyl)-4-methylphenol (BHT; 2) have been glycosylated by thermal reaction (20-60°) with various glycosylidene-derived diazirines.4-Methoxyphenol reacted with the D-glucosylidene-derived diazirine 3 to give O-glucosides (4 and 5, 69%, 3:1) and C-glucosides (6 and 7, 16%, 1:1). Similarly, phenol yielded O-glucosides (10 and 11, 70%, 4:1) and C-glucosides (12 and 13, 13%, 1:1). 4-Nitrophenol gave only O-glycosides, 3 leading to 14 and 15 (75%, 3:2; Scheme 1), and the D-galactosylidene-derived diazirine 17 to 22 and 23 (52% (from 16), 65:35; Scheme 2). The reaction of phenol with 17 yielded 58% (from 16) of the O-galactosides 18 and 19 (4:1) and 14% of the C-galactosides 20 and 21 (1:1). From the D-mannosylidene-derived diazirine 25, we predominantly obtained the α-D-configurated 26 (38 % from 24). These results are interpreted by assuming that an intermediate (presumably a glycosylidene carbene) first deprotonates the phenol to generate an ion pair which combines to give O- and - with electron-rich phenolates - also C-glycosides. A competition experiment of 3 with 4-nitro- and 4-methoxyphenol gave the products from the former (14 and 15) and the latter phenol (4-7) in almost equal amounts. Differences in the kinetic acidity of OH groups, however, may form the basis of a regioselective glycosidation, as evidenced by the reaction of 3 with methyl orsellinate (1) yielding exclusively the 4-O-monoglycosylated products 27 and 28 (78%, 85:15), although diglycosidation is possible (27→ 31 and 32; 67%, 4:3; Scheme 3). Steric hindrance does not affect this type of glycosidation; 3 reacted with the hindered BHT (2) to afford 33 and 34 (81 %, 4:1). The predominant formation of 1,2-trans -configurated O-aryl glycosides is rationalized by a neighbouring-group participation of the 2-benzyloxy group.

Notes: The fundamental molecular aspects of trialkyltin compounds of the type R3SnY have been investigated in view of their applicability as ion-selective components in solvent polymeric membranes. The interaction between these compounds and anions has been studied using 119Sn- and 13C-NMR. Neutral tetracoordinated trialkyltin compounds form a negatively charged pentacoordinated complex upon interaction with Cl-ions in homogeneous organic phases as well as in membranes in contact with aqueous solutions. Although in a homogeneous phase, the electronegative substituent Y determines the complex-formation constant, it has no influence on the potentiometric anion selectivity in liquid membranes containing trialkyltin carriers R3SnY with different Y. The observed selectivity pattern is not given by the magnitude of the stability constants in a homogeneous phase but is dictated by the prevailing association-dissociation process leading to tetracoordinated compounds which change in constitution due to varying sample composition, The results obtained from equilibrium studies of tetravalent mono tin compounds with anions in both homogeneous phase and in two-phase systems confirm the earlier hypothesis that trialkyltin compounds incorporated in solvent polymeric membranes act as electrically neutral carriers for anions.

Notes: High-level ab initio calculations have been made for fluoromethylamine to study structural and energetic effects of the relative orientation of the N lone pair to the C—F bond. The anti-conformer (N lone pair anti-planar to the C—F bond) corresponds to the global energy minimum. It has the longest C—F distance, the shortest C—N distance, and is 7.5 kcal·mol-1 more stable than the related perpendicular conformation (lone pair perpendicular to the C—F bond). The syn-conformation also shows hallmarks of the anomeric effect: long C—F bond, short C—N bond, and energetic stability when allowance is made for the two pairs of eclipsed hydrogens. The transition state for N inversion is close to the syn-structure; rotation about the C—N bond is strongly coupled with this inversion process. Small bond distance changes of ca. 0.02 Å between parallel and perpendicular conformations are associated with dissociation energy differences of ca. 30 kcal·mol-1. Various criteria for assessing the strength of the anomeric effect are discussed.

Notes: Flavonoid disaccharide monoglycosides have been acylated by the catalytic action of the protease subtilisin in anhydrous pyridine. The effects of the nature of the sugars and of the interglycosidic bonds on the regioselectivity of the reactions have been analyzed. The selectivity was excellent with rutin (1), hesperidin (2), naringin (6), and quercetin 3-O-[O-(β-D-glucopyranosyl)-(1→4)-α-L-rhamnoside] (9), giving single monoesters on their glucose moieties (see la, 2a, 6a, and 9b, resp.); quite interestingly, in the last compound, acylation did not occur at the free primary OH group but at the secondary OH—C(3‴). On the other hand, a mixture of mono- and diesters was obtained with the flavonoid peltatoside (7).

Notes: 4-Hydroxycoumarin (= 4-hydroxy-2H-1-benzopyran-2-one) reacts with enals to give 1,2- or 1,4-addition products, depending on the nature and relative location of the substituents on the olefinic double bond (Scheme 2). The resulting adducts further react intra- or intermolecularly, affording dimeric coumarins or pyranocoumarins in the case of 1,2-addition and acetalic pyranocoumarins in the case of 1,4-addition. With enals bearing alkyl groups at C(β), 2H-pyrano[3,2-c]coumarins are the only products formed, and the reaction represents an easy and straightforward entry into this class of recently described biologically active natural products.

Notes: The synthesis of several configurationally defined hydroxymilbemycin derivatives is described. One of these allylic alcohols is the known 5-O-[(tert-butyl)dimethylsilyl]-13α-hydroxymilbemycin D (= 5-O-[(tert-butyl)-dimethylsilyl]-22,23-dihydroavermectin B1b, aglycone; 15D), the synthesis of which represents LI conversion of the milbemycin to the avermectin series of natural products. The configurations at C(13), C(14), and C(15) of the new milbemycin derivatives were determined by NMR experiments and force-field calculations.

Notes: The four possible A/B cis-fused diastereoisomers of Ambrox® have been synthesized and their configurations and conformations established by X-ray and NMR analysis. Only 5β-ambrox (= 1,2,3a,4,5,5β,6,7,8,9,9a,9bα-dodecahydro-3aβ,6,6,9aβ-tetramethylnaphtho[2,1-b]furan; 5) has an odor quality comparable to Ambrox®. The 1,3-synperiplanar/diaxial conformation of the substituents at C(8) ( = C(3a)) and C(10) (= C(9a)) has thus been confirmed to be a compulsory structure element for the particular odor.

Notes: The three ligands 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetrakis(methylenephosphonic acid) (1), 1,4,7,11-tetraazacyclotridecane-1,4,7,11-tetrakis(methylenephosphonic acid) (2), and 1,4,8,11-tetraazacyclotetradecane-1,4,8,1 1-tetrakis(methylenephosphonic acid) (3) have been synthesized by condensation of the corresponding macrocycles with formaldehyde and phosphorous acid. The protonation and stability constants with the earth-alkali ions have been determined at 25° and I = 0.1 M (Me4)N(NO3) by potentiometric titrations. Because of the high values of the first two protonation constants, 1H-NMR measurements were necessary to determine them. Titrations in different supporting electrolytes (NaNO3, KNO3, RbNO3, CsNO3, and Me4N(NO3)) show that their choice is of paramount importance, as the above ligands can form complexes with alkali-metal ions. The potentiometric results for the earth-alkali ions show that beside mononuclear complexes of different degrees of protonation ([MLHn], n = 0-4), also binuclear species are formed ([M2LHm], m = 0-2). It is interesting that 1 with the smallest macrocyclic ring has the greatest tendency to form binuclear complexes, which are so stable that they partially prevent the formation of the corresponding mononuclear species. For [ML], [MLH], [M2L], and [M2LH], the stability sequence is Mg2+ 〈 Ca2+ 〉 Sr2+ 〉 Ba2+, whereas for [MLH2], [MLH3], and [MLH4], the stability steadily decreases from Mg2+ to Ba2+.

Notes: The title complex crystallises in two C3v, isomeric forms differing in carbonyl-ligand arrangement. In solution, the isomer 1b with three edge-bridging carbonyls on a common face of the metal tetrahedron converts via an endothermic equilibrium into the isomer 1u with no bridging carbonyls. The latter was shown by 13C-NMR to be the intermediate of the ‘merry-go-round’ process which exchanges the sites of the basal CO's.

Notes: The ‘naked sugar’ (+)-(1R,2R4R)-2-endo-cyano-7-oxabicyclo[2.2.1]hept-5-sn-2-exo-yl acetate ((+)-4) was converted (7 steps, 45% overall) with high stereoselectivity into (-)-(4R,5S,6R)-4,5,6-tris{[(tert-butyl)dimethylsilyl]oxy}cyclohex-2-en-1-one ((-)-11). Reduction of (-)-1 with NaBH4- CeCl3 · 7 H2O, followed by deprotection of the silyl ether moieties gave (+)-conduritol F ((+)-1; 47%) whose characteristics were identical to those of natural (+)-leucanthemitol. Reduction of (-)-11 with DIBAH, followed by deprotection of the silyl ether moiety led to (-)-conduritol B ((-)-3; 51 %).

Notes: Adamantanone-derived nitrone 4 and some other keto-nitrones, when reacted with aromatic and aliphatic aldehydes in refluxing toluene or tetrahydrofuran, formed the corresponding aldonitrones (Z)-10, the latter arising from the fragmentation of an initially formed 1,4,2-dioxazolidine 6 to adamantan-2-one and an oxaziridine intermediate 11, which then rearranges to (Z)-10.

Notes: The synthesis of some derivatives of the title compound VI is described. Bromination of diethyl (cis-3-azido-2-oxo-4-styrylazetidin-1-yl)(pyridin-2-yl)methylphosphonate (6) in MeOH gave tricyclic β-lactam 7, while similar bromination of diethyl (cis-3-azido-2-oxo-4-vinylazetidin-1-yl)(pyridin-2-yl)methylphosphonate (9) afforded tri-cyclic β-lactam 10. Mechanisms for these transformations are proposed (Schemes 1 and 2).

Notes: A new vicinal dioxime ligand with two crown-ether groups, 1,2-bis[(monoaza[15]crown-5)-N-Yl]-glyoxime(LH2), has been prepared from cyanogen di-N-oxide and monoaza[15]crown-5. Ni(II), Pd(II), and Pt(IV) complexes of LH2 with or without alkali-metal ions bound to macrocyclic groups have been isolated. The high affinity of [Pd(LH)2] and [Ni(LH)2] for the K+ ion is observed in solvent extraction experiments. A single-crystal X-ray structure confirms the postulated geometry of [Pd(LH)2]- The Pd-atom of the centro-symmetric molecule has square-planar PdN4 coordination where Pd-N distances range from 1.978(3) to 1.970(3) Å. The N-Pd-N intraligand angle is 79.9(1)°.

Notes: The protected intermediate (11R)-25-O-deacetyl-11-deoxo-11-hydroxy-21,23-O-isopropylidenerifamycin S (7) has been synthesized starting from rifamicin S (2; Scheme 2), the former being a potential substrate for the preparation of new types of rifamicin-S derivatives modified at C(11) and/or C(25). The reactivity of 7 toward acylations has been studied under both base- and acid-catalyzed conditions. The compound either did not react or nr underwent unexpected reactions, and no acylation products could be isolated. The X-ray crystal structure of 7 reveals that both OH groups at C(11) and C(25) are hindered, and this is probably the reason, why other take place faster than acylation.

Notes: In chloroform, [ZrCl4·2(MeO)3PO] exists in both cis- and trans-isomeric forms. Three reactions can be envisaged in the presence of excess (MeO)3PO = L: (1) cis-[ZrCl4·2L] + *L⇆cis-[ZrCl4·L*L]+ L; (2) trans-[ZrCl4·2L] + *L ⇆ trans-[ZrCl4·L*L] + L; (3) cis-[ZrCl4·2L]⇆ trans-[ZrCl4·2L]. To distinguish between these possible reaction pathways, we have used 2D 1H-NMR spectroscopy. For the first time, variable-pressure 2D exchange spectra were used for mechanistic assignments. cis/trans-Isomerisation was found to be the fastest reaction (in CHCl3/CDCl3), with a small acceleration at higher pressure: it is concluded to be an intramolecular process with a slightly contracted six-coordinate transition state. The intermolecular (MeO)3PO exchange on the cis- and trans-isomer are second-order processes and are strongly accelerated by increased pressure: Ia mechanisms are suggested without ruling out limiting A mechanisms.

Notes: For certain ionophores of extremely high lipophilicity, kinetic limitations of the carrier-induced ion transfer between aqueous and membrane phase may heavily disturb the electromotive behaviour of the ion-selective membrane electrode. These limitations may be overcome by adding tetraphenylborates to the membrane phase. Membranes prepared with different 3,6-dioxaoctanediamide homologues and potassium tetrakis(P-chlorophenyl)borate as phase-transfer catalyst all exhibit the same ion selectivity as well as theoretical electrode response. This behaviour is corroborated by a theoretical description of the ion-transfer process.

Notes: Solutions of blockcopolymers (POE-b-PI-b-POE) in fluids of interacting aqueous nanodroplets (W/O microemulsions) are studied. The interaction strength between the (pseudo) two components is measured by the shift of the percolation temperature relative to that of the pure microemulsion. A quantitative measure of the interaction, the differential heat of solution, is thermodynamically related to the slopes of the equilibrium temperature of the system with varying monomeric nanodroplet concentration and the experimental percolation line in the presence of copolymer.

Notes: Synthesis of Cyclic Depsipeptides via Direct Amide Cyclization: Cyclic Depsipeptides with 12-Ring Atoms and Alternating Sequence of α-Hydroxy and α-Amino AddsThe reaction of 3-(dimethylamino)-2,2-dimethyl-2H-azirine (1; R1 = R2 = R3 = R4 = Me) with α-hydroxy-carboxylic acids, followed by selective hydrolysis of the terminal dimethylamide group yields the dipeptide analogues 15a and 18b (Schemes 3 and 4). After protection of the OH group (→ 16a and 19, resp.), coupling with the C-terminus-protected derivatives 14 and 18a, respectively, by a modified 1,1′-carbonyldiimidazole procedure followed by hydrolysis gives the linear depsipeptides 17c and 20, respectively. Treatment with HCl gas in toluene at 100° leads to the cyclic depsipeptides 21 and 22 in very good yield. The two model reactions show that the ‘azirine/oxazolone-method’, combined with the ‘direct amide cyclization’, is a versatile procedure for the synthesis of cyclic depsipeptides containing α,α-disubstituted α-amino acids.