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Notes: We have calculated the translational and rotational frictional coefficients of structures related to T2 and T4 bacteriophage, using the theoretical framework developed in the preceding two papers. The structures considered were models for tail-fiberless phage, and for whole phage with fibers in the extended and retracted portions. We also computed and compared with the experiment the changes in translational frictional coefficient produced by successive addition of 1-6 fibers to the fiberless particle. Agreement with experimental results is markedly improved over previous theoretical efforts, especially with respect to the effect of tail-fiber extension. Some significant discrepancies remain, however, in the comparison of fiber-retracted and fiberless phage.

Notes: We have used the modified Oseen hydrodynamic interaction tensor along with iterative numerical solution of the coupled hydrodynamic interaction equations to calculate the rotational diffusion coefficients of macromolecular complexes composed of nonidentical spherical subunits. For the one structure, a prolate ellipsoid of revolution, for which exact solutions are available, a subunit model with the same length and volume gives asymptotic agreement with the Perrin equations. Other structures considered include plane polygonal rings, lollipops, and dumbbells.

Notes: We have studied the effects of temperature, EDTA, and ionic strength on C-polysaccharides in solution by examining the details of the time-correlation function using a 96-channel single-clipped photon correlation spectrometer. Our linewidth results have shown that the C-polysaccharides in buffer solution form aggregates of very broad distributions. Thus, fractionation by gel-filtration chromatography is only mildly effective. Although the aggregate sizes seem to remain relatively constant from 4 to 25°C, a fraction of those aggregates break up to form smaller fragments or monomers at higher temperatures. However, the dissolution-association process is quite slow and takes days even at room temperatures before the equilibrium is reached. We have also shown that by adding an excess amoutn of EDTA, the aggregates can be broken up. Again the dramatic changes occur only at short delay times suggesting that a protion of the larger aggregates remains. Finally, the amount and size of aggregates depend upon the ionic strength which exhibit a maximum ΓT/sin2 (θ/2) around 0.1-0.2 M KCl.If the activities of polysaccharides in solution depend upon molecular size, the standard techniques such as gel-permeation chromatography and ultracentrifugation cannot properly characterize the detailed size distribution. Quasielastic laser light scattering can provide us with a qualitative model. The quantitative details must necessarily await more extensive investigations using a combination of the techniques and better fractionation procedures in an appropriate buffer solution.

Notes: The coupling of N-acyl-α-amino-acids with α-hydroxyacid-methyl amides results in depsipeptide molecules containing two chiral centers and one ester function inserted between two amide functions. Their conformational features have been investigated by IR spectroscopy, proton magnetic resonance, X-ray diffraction, and theoretical P.C.I.L.O. calculations.It is shown that most of these molecules are folded by an intramolecular 4 → 1 hydrogen bonding. Two folded conformations, similar to the well known β turn in peptides, are described, the stability of which depends on the configurational sequence in the investigated molecule.LL and LD species are folded in two different ways whereas LG sequences containing an achiral hydroxy-acid residue accommodate both of them. The presence of a N-terminal achiral amino acid noticeably decreases the folding ratio.The above conclusions are then compared with the conformational features of homologous tripeptide molecules.

Notes: The kinetics of complex formation between fluorescein mercuric acetate and heat-denatured DNA were studied by measuring the fluorescence quenching of this reagent. This quenching process involved no immeasurably rapid phase and it was shown that this reaction follows simple second-order kinetics. The rate constant at 25°C was estimated to be 2.9 × 104M-1 sec-1 for calf-thymus DNA (42% G + C) and 1.1 × 104M-1 sec-1 for Micrococcus lysodeikticus DNA (72% G + C). Activation parameters for this reaction were calculated from the temperature dependence of the reaction rate, and the activation entropy was found to be highly negative (-27.5 cal/mol deg for calf-thymus DNA and -25.5 cal/mol deg for M. lysodeikticus DNA). The binding of fluorescein mercuric acetate to native DNA, which requires the opening of the double-helical structure, was also followed by measuring the absorbance change of this reagent. There was a lag phase in this binding process, and the enthalpy change for the opening step corresponded roughly to that for the opening of one base pair. These findings are discussed in relation to the results of a similar study with formaldehyde.

Notes: The formation of collagen fibrils from soluble monomers and aggregates by thermal gelation at neutral pH can be divided into two distinct stages: a nucleation phase and a growth phase. Turbidity studies of the kinetics of the precipitation reaction show that the lag-phase time or nucleation reaction time, t′l, is markedly temperature dependent while the growth reaction time is temperature independent. The activation energy of the nucleation reaction is essentially constant over the temperature range studied. In monitoring the nucleation-phase reaction by various physicochemical techniques, including viscosity, sedimentation equilibrium, and light scattering, no evidence for the formation of aggregates was observed. Enrichment of the initial collagen solution with aggregates accelerates nucleation, but de novo nuclei formation is still required even in highly aggregated collagen preparations. Removal of pepsin and pronase susceptible peptides lengthens the nucleation reaction time and increases the sensitivity of the rate of nuclei formation to changes in ionic strength. Electron microscope studies show the fibrils formed from the protease-treated collagen to be less well organized. With pepsin-treated collagen, subfibrils and obliquely striated fibrils are seen, showing that while microfibrils are formed interactions between them are modulated by the enzyme susceptible peptides in the same way that these regions modulate nuclei assembly. It appears that pepsin and pronase susceptible peptide regions of collagen play a more prominent role in the in vitro assembly of collagen molecules to form D-stagger nuclei and fibrils than do ionic interactions between helical molecular regions. A mechanism of nucleation of collagen fibrillogenesis is discussed.

Notes: Circular dichroism spectra of the nucleic acid monomers have been measured in aqueous solution and extended into the vacuum ultraviolet region to about 166 nm. Measurements were made on ribo and deoxyribo derivatives of adenine, guanine, hypoxanthine, cytosine, thymine, and uracil derivatives both with and without the 5′-phosphate (with the exception of ribosyl thymine 5′-phosphate). Absorption spectra of the deoxyribonucleotides measured to about 175 nm are also presented. The results demonstrate that both the circular dichroism and absorption spectra observed below 200 nm are no more complicated than the spectra normally recorded above 200 nm. In most cases, the circular dichroism spectra of the various derivatives of a given base are similar, indicating that the conformations are similar. On the other hand, the differences among the circular dichroism spectra of the various derivatives of a given base are sufficient to identify a particular derivative. The average circular dichroism for the deoxyribonucleotides is compared with the circular dichroism of native E. coli DNA. The comparison reveals that the circular dichroism of DNA below 200 nm is due principally to the interaction between the bases rather than the intrinsic circular dichroism of the monomers. The monomer transitions are discussed in relationship to the absorption and circular dichroism spectra presented.

Notes: A theory for the determination of DNA arrangements in DNA-containing specimens, using planar aromatic dye molecules as probes for plane polarization of fluorescence, has been described. At low dye-to-DNA concentrations, the dye molecules are sandwiched between the stacked bases of DNA; hence, the fluorescence from the dye bound to a local region of DNA helix is plane-polarized with the polarization direction perpendicular to the local axis of DNA. The degree of such polarization from an aligned DNA-specimen complexed with dye is determined both by the DNA orientation and the conformational state (e.g., base tilt) of DNA into that specimen. Analysis has been made of the relationship between the degree of polarization and the orientation of the emitting dipoles of dye. The dye complexes may be aligned in a mechanical shear or electric field. However, any change in the orientation distribution of the emitting dipoles due to force fields should be taken into account. With some assumptions and approximations, the magnitude and the direction of maximum polarization can be related to different orders of DNA coiling and to their various combinations. Since the measured polarization is averaged over all DNA regions of the specimen, if the magnitude of polarization is appreciable and the polarization occurs in the specific direction of the specimen, the theory helps to eliminate several probable arrangements of DNA. The predominant molecular features of the actual DNA arrangement can be determined through this process of elimination, as explained in two subsequent papers with T-even bacteriophage and chromosome systems.

Notes: Fluorescence of 2-p-toluidinylnaphthalene-6-sulfonate (TNS) was enhanced in the presence of maltooligosaccharides, amylose, and other α-glucans. The dependence of relative TNS fluorescence intensity per glucose unit on chain length of oligosaccharides was examined. The values of binding constant and thermodynamic parameters, assuming the 1:1 complex for TNS-amylose (number-average degree of polymerization, DPN = 17), were determined by the fluorescence titration. The values of thermodynamic parameters for 1:1 complex formation of TNS-α- and β-cyclodextrins were also determined and compared with those of TNS-amylose (DPN = 17). The fluorescence intensity of TNS in the presence of amylose (DPN = 600) decreased by the action of glucoamylase and taka-amylase A. The fluorescence of TNS-amylose (DPN = 17) system increased with the increased ionic strength. In the presence of pullulan, TNS fluorescence was also enhanced and decreased by the action of pullulanase. Amylopectin enhanced TNS fluorescence rather more strongly than amylose (DPN = 17) at the same concentration. In the presence of dextran, the fluorescence of TNS was scarcely enhanced. The degree of fluorescence enhancement of TNS in the presence of α-glucans seems to reflect the structures of α-glucans in solution, since TNS fluorescence is enhanced in the hydrophobic environment or by the disturbance of free intramolecular rotation.

Notes: Water-soluble, random copolymers containing L-methionine and N5-(3-hydroxypropyl)-L-glutamine have been prepared, fractionated, and characterized. The thermally induced helix-coil transitions of these copolymers in water have been investigated, and it has been found that incorporation of L-methionine increases the helix content of the polymers at all temperatures in the range of 0-60°C. The Zimm-Bragg parameters σ and s for the helix-coil transition in poly(L-methionine) in water were deduced from an analysis of the melting curves of the copolymers using the methods described in earlier papers.

Notes: Normal vibrational analysis was carried out for DNA molecules in both A and B conformations as well as for A-RNA. A simplified backbone model was examined and expanded to include the backbone phosphate-group and the ribose ring. We applied the new force-constant refinement procedure discussed in the preceeding paper [Van Zandt, L. L., Lu, K.-C. & Prohofsky, E. W. (1977) Biopolymers, 16, 2481-90] to fit some observed frequencies in the Raman spectra for all three nucleic acids with the same set of force constants. The results indicate that the observed frequency shift can be attributed to the conformational change solely. We ignored the second-order differences in force constants for the different geometries. The agreement between the observed and calculated frequencies derived from the final refined set of force constants is good and apparently justifies this assumption. Two modes previously assigned to the symmetric diester O-P-O stretch and the symmetric dioxy O‥P‥O stretch are actually fitted. They are mainly backbone phosphate-group modes. The refined ribose-ring force-constants were transferred to the calculation of the vibrational spectrum of tetrahydrofuran. The overall agreement is again good. We discuss these calculations and the resulting normal modes. We also discuss the application of the Green-function refinement scheme and several strategies adopted to bias the convergence of the procedure.

Notes: A study of the conformational states of the dinucleotide coenzyme NAD+ has been made using semiempirical energy calculations. Taking low-energy mononucleotide structures as starting conformations, energy minimizations have been performed. The lowest energy states are stacked structures, with interactions between the adenine and nicotinamide rings. Some structures show stabilization gained from electrostatic attractions between the positively charged nicotinamide and negatively charged phosphate oxygens. These predictions correlate well with the available experimental data.

Notes: Water-soluble block copolymers of the type (A)m-(B)n-(A)p, where (A)m,p was either poly(D,L-lysine-α,β,β,γ,γ,δ,δ-d7) or poly(D,L-lysine) and (B)n was either poly(L-alanine) or poly(L-phenylalanine), were synthesized for conformational studies by proton magnetic resonance spectroscopy. Analytical determination of the amount of the initiator fragment (n-hexylamine) at the C-terminus of the copolymers was used to obtain the number-average degrees of polymerization, DPn, and thereby, together with the amino acid composition, to establish the covalent structures of the polymers. The values of DPn were found to be much lower than those deduced from sedimentation equilibrium or form viscosity measurements. These deviations, which also are thought to have arisen in similar studies reported in the literature, are attributable to intermolecular aggregation; the relation of such aggregation to covalent structure (and its effect on the polymerization reaction) is discussed in terms of the conditions and mechanism of synthesis of block copolymers of amino acids.

Notes: Poly(Lys48, His52), a random copolypeptide of L-lysine (48%) and L-histidine (52%), was used as a model protein for investigating the effects of protonation on the imidazole group of histidines on protein binding to DNA. The complexes formed between poly(Lys48, His52) and DNA were examined using absorbance, circular dichroism (CD), and thermal denaturation. Although increasing pH reduces the charges on histidine side chains in the model protein, the protein still binds the DNA with approximately one positive charge per negative charge in protein-bound regions. Nevertheless, CD and melting properties of poly(Lys48, His52)-DNA complexes still depend upon the solution pH which determines the protonation state of imidazole group of histidine side chains. At pH 7.0, the complexes show two characteristic melting bands with a tm (46-51°C) for free base pairs and a t′m (94°C) for protein-bound base pairs. The t′m of the complexes is reduced to 90°C at pH 9.2, although at this pH there is still one lysine per phosphate in protein-bound regions. Presumably, the presence of deprotonated histidine residues destabilizes the native structure of protein-bound DNA. The binding of this model protein to DNA causes a red shift of the crossover point and both a red shift and a reduction of the positive CD band of DNA near 275 nm. This phenomenon is similar to that caused by polylysine binding. These effects, however, are greatly diminished when histidine side chains in the model protein are deprotonated. The structure of already formed poly(Lys48, His52)·DNA complexes can be perturbed by changing the solution pH. However, the results suggest a readjustment of the complex to accommodate charge interactions rather than a full dissociation of the complex followed by reassociation between the model protein and DNA.

Notes: A theoretical analysis of the conformation of the octapeptide hormone Asn1, Val5 angiotensin II has been carried out by semiempirical potential energy calculations. A preliminary study of the Ala6-Pro-Ala molecule, which mimics the angiotensin backbone, provided us with likely backbone structures on which the effect of the full side chains of the hormone could be assessed. For angiotensin II, the calculations show that only a small number of folded, compact conformations have a high probability of existence. This is the consequence of favorable packing and of the presence of proline in position 7. These results are consistent with various experimental data, both structural and biological. This method is readily applicable to the study of analogs of the hormone or to other peptides of comparable size.

Notes: The interaction between DNA and ionen polymers, -[N+(CH3)2(CH2)mN+(CH3)2(CH2)n]l-, with m-n of 3-3, 6-6, and 6-10 were examined in order to know how the binding behavior of cationic polymers with DNA depends on the charge density of polycation. The ionen polymer has no bulky side chain and the binding forces with DNA would be attributed mainly to electrostatic interaction. When 3-3 ionen polymers were added to DNA solution, precipitable complexes with the ratio of cationic residue to DNA phosphate (+/-) of 1/1 and the free DNA molecules were segregated, while 6-6 and 6-10 ionen polymers formed soluble complexes with DNA molecules up to (+/-) = 0.5. This suggests that 3-3 ionen polymers bind cooperatively with DNA while 6-6 and 6-10 ionen polymers bind noncooperatively. The cooperative binding of 3-3 ionen polymer and the noncooperative binding of 6-6 ionen polymer were also supported by the thermal melting and recooling profiles from the midpoint between first and second meltings. It was concluded that the charge density of DNA phosphate is a critical value determining whether the ionen polymers bind to DNA by a cooperative or by a noncooperative binding, since the distance between successive cationic charges of 3-3 ionen polymer is shorter than that between successive phosphate charges on DNA double helix and those of 6-6 and 6-10 ionen polymers are longer.

Notes: Quasielastic light scattering is used to study the effect of ionic strength on the dynamic behaviour of DNA. In a first approach the spectrum of scattered light is analyzed in terms of a single relaxation process. The large difference between the observed behaviour and that expected according to a pure diffusional process reflects the contribution associated with internal modes, which increases with decreasing ionic strength. Such behaviour is better analyzed in terms of a double relaxation process by using two relaxation times, the reciprocals of which are equal to DK2 and DK2 + τi-1 (K), respectively, where τi (K) is an average value describing the set of modes observed at a given K value. Relative intensity and relaxation times, which are the more accurate parameters, were used to interpret the results. The observed increase of the relative contribution of internal modes with decreasing ionic strength is actually a relative decrease of the diffusional contribution induced by a corresponding increase of the radius of gyration RG. On the other hand, the reciprocal τi-1 (K) of the relaxation time is a linear function of K2 in the analyzed KRG range and is insensitive to ionic strength between 10-2M and 1M. These results, when discussed according to Rouse's model, lead to define for each value of τi-1 (K) a corresponding mean-squared equilibrium length 〈μi2〉 which is found to be a linear function of K-2.

Notes: Derivatives and peptides of β-nitrobenzyl-L-aspartates were studied with high-field nmr. Differences were observed between the chemical shifts of protons located near the extremity of the principal chain as a function of the terminal group. These differences are explained by conformational calculations which exclude the existence of an hydrogen bond and demonstrate the influence of the aromatic ring position on the protons of the main chain. Both nmr experiments and conformational analysis indicate that conformations are nearly the same for ortho, meta, and para nitro substitution. These conclusions are in good agreement with Karplus relationship applied to the α and β protons.

Notes: Excess small angle X-ray scattering in solvents of differing electron density has been calculated from the crystal structures obtained for rubredoxin, trypsin inhibitor, myogen, ferricytochrome c2, ribonuclease S, lysozyme, nuclease, myoglobin, α-chymotrypsin, elastase, subtilisin, carboxypeptidase A, thermolysin, methemoglobin, deoxyhemoglobin, and a single polypeptide chain of M4 lactate dehydrogenase. The scattering curves for each protein can be reproduced by the sum of three curves, with the weighting of the three curves depending on the electron density of the solvent. The radius of gyration obtained from the small angle X-ray scattering by globular proteins in aqueous solution will usually exceed the values defined by the shape of the macromolecule. Deviations for certain of the proteins cited are calculated to be as large as 6%. These deviations arise from the tendency for the amino acid residues with low electron density to be situated closer to the center of the protein than the amino acid residues of high electron density. An upper limit of 19% is obtained for the discrepancy between the radius of gyration defined by the shape of a spherical globular protein of typical amino acid composition and the apparent radius of gyration measured for that protein in water by small angle X-ray scattering.

Notes: The ability of the macrotetrolide nactins to complex selectivity with a wide variety of cations makes these ionophorous antibiotics important model systems for the study of biologic ionic transport. We report a Raman spectroscopic investigation of the Na+, K+, Rb+, Cs+, Tl+, NH4+, NH3OH+, C(NH2)3+, and Ba++ complexes of nonactin, monactin, and dinactin in 4:1 (v/v) CH3OH/CHCl3 and in the solid state. The nactins display characteristic spectral changes upon complexation, some of which are specific for a given cation. In the K+, Rb+, Cs+, NH3OH+, and C(NH2)3+ complexes, which are apparently isosteric, the ester carbonyl stretch frequency is found to be linearly proportional to the cation-carbonyl electrostatic interaction energy, as calculated from a simplified model. Deviations for the Na+, NH4+, Tl+, and Ba++ complexes are interpreted as arising from additional nonelectrostatic interactions. Additional information is obtained from other spectral regions and from measurements of depolarization ratios. Spectra of the nactin complexes differ from each other more in the solid state than in solution, reflecting the effects of crystalline contact forces.

Notes: The interaction of water with collagenous tissue was investigated using dynamic mechanical spectroscopy and cryogenic X-ray techniques. The loss spectrum was found to be very sensitive to water which is highly associated with the macromolecule. Two water-sensitive loss peaks were observed below 0°C: the β2 or “water dispersion” at 150°K and the β1 at 200°K which is attributed to motion of polar side chains. Changes in peak temperature and intensity were not continuous with water content, but exhibited regimes in behavior which were associated with two types of nonfreezable water, structural and bound water. In cryogenic X-ray experiments, specimens which contained some freezable water exhibited reflections identified with the cubic form of ice. These ice crystals underwent an irreversible transition to the more common hexagonal form when warmed above 200°K. On the basis of these experiments, a model for the hydration of native collagenous tissue was proposed.

Notes: Conformation behavior of polyethylenimine has been examined by studies of the fluorescence characteristics of derivatives of the polymer containing pyrenyl ligands. Excimer formation within the macromolecular matrix serves as a sensitive probe of group proximities. The experimental observations combined with nearest neighbor analyses lead to quantitative assessments of the extent of interaction between polymer side chains.

Notes: The chiroptical properties of S-proline conformational isomers are examined on a theoretical model in which electronic wave functions are obtained from semiempirical molecular orbital calculations. The CNDO/S molecular orbital model is used to perform SCF-MO calculations on ground state electronic structure and excited states are constructed in the virtual orbital-configuration interaction approximation. Electronic rotatory strengths and dipole strengths are calculated directly from the complete (but approximate) molecular electronic wave functions. Zwitterionic, cationic, and anionic S-proline structures are studied twotypes of conformational variables are represented in the calculations: (1) pyrrolidine ring conformation; and (2) rotation about the Cα-COO- bond. Rotatory strengths are found to be somewhat sensitive to rotational isomerism about the Cα-COO- bond, but are found to be rather insensitive to conformational changes within the pyrrolidine ring. The CD spectrum of zwitterionic S-proline down to ∼160 nm appears to be well accounted for by the theoretically calculated results if conformational preferences with respect to rotation about the Cα-COO- bond can be assumed to exist in solution media. Furthermore, spectra-structure correlations are offered for the anionic and cationic forms of S-proline in solution.

Notes: A series of stereoregular α-(1 → 6) linked glucomannans have been prepared by Lewis acid-catalyzed copolymerization of anhydro sugar derivatives followed by debenzylation. The products have been characterized for mole fraction of the individual monomer, and sequence lengths have been calculated from copolymerization data. The viscosity, specific rotation, and 13C nmr spectra have been correlated with the structure of the various copolymers.

Notes: This paper presents the results of a stereochemical analysis of local interactions in unfolded protein chains (sterical repulsions, hydrogen, and hydrophobic bonds, etc.) by means of space-filling modeles. On the basis of this analysis, an evaluation is made of thermodynamic parameters controlling the building-in of all the 20 natural amino acid residues in all the physically possible position of local secondary structures (α-helices, including α-helices with short fragments of helices 310 at the C-terminus; β-bends of different types, helices 310, and their combinations) as well as thermodynamic parameters of separate hydrogen bonds of polar side groups with the neighbor peptide groups (“local contacts”). The accuracy of the obtained results is discussed.

Notes: The translational diffusion coefficient of a pure sample of α-chymotrypsinogen A is measured by laser light scattering to give a value of D20,w0 = (8.40 ± 0.15) × 10-7 cm2/sec.

Notes: The exchange rate of the indole nitrogen proton with solvent water protons was measured as a function of pH and temperature for tryptophan, N-methyl tryptophan, glycyl tryptophan, tryptophanamide, tryptophylglycine, and tryptophylglycyl glycine. The nmr observation was by long-pulse Fourier transform methods, and kinetics were inferred from saturation recovery, H2O transfer of saturation, and linewidth. There are observable differences between the rates of these compounds, but all are describable within a factor of two by specific acid and base-catalyzed rates kH = 100 and kOH = 108 1/mol-sec at 27°C. It is concluded that this behaviour is representative of this proton on the indole side chain in a random-chain peptide exposed to water.

Notes: Stoichiometric mixtures of acridine orange with dermatan sulfate at total dye concentrations ≤ 1 × 10-5 M show fluorescence maxima at 540 nm and 660 nm on excitation at 436 nm. By means of membrane filtration, it is directly demonstrated that the species emitting at 540 nm is due only to unbound dye whereas the 660-nm emitting species is due to bound dye. It is, therefore, possible to differentiate unbound acridine orange from its dermatan sulfate complex solely by spectroscopic methods. Thermodynamic binding parameters can be calculated from rapid spectroscopic measurements without disturbing the system.

Notes: A general kinetic analysis for unimolecular three-species models is presented. Criteria are established for each model in order to allow selection among alternative models on the basis of their amplitude behavior. These criteria are applied to the previously reported kinetic data for the folding transitions of lysozyme and cytochrome c in guanidine hydrochloride. The degree of success in applying these criteria is found to depend on the extent to which the apparent rate constants, λ1 and λ2, approach each other in certain conditions. Ikai and Tanford have presented a similar kinetic analysis, although these authors considered only the case where no intermediate kinetic species was populated initially. For one protein system, namely, ribonuclease A (RNase A), at least one stable intermediate exists after unfolding outside of the equilibrium transition zone. The criteria developed by Ikai and Tanford cannot be applied to RNase A kinetics and, therefore, a more general analysis is presented. Previously published studies of the reversible folding kinetics of lysozyme and cytochrome c are reexamined using the current amplitude analysis, and it is found that both of these folding transitions can be described by the three-species model, \documentclass{article}\pagestyle{empty}\begin{document}$$ U_{1} \rightleftharpoons U \rightleftharpoons N $$\end{document} used previously to describe the folding kinetics of RNase A (U1 and U2 are two forms of unfolded enzyme and N is the native species). These results suggest that this three-species model may provide a fairly general description of the reversible folding kinetics of small proteins. As an outgrowth of the current analysis, several suggestions for experimental design in studying protein folding will be presented and discussed.

Notes: The copolymer which has both ligand sites (4-vinylpyridine) and redox sites (N-(p-vinylbenzyl)-3-carbamoyl-1,4-dihydropyridine) was synthesized by the dithionite reduction of the copoly(4-vinylpyridine-N-(p-vinylbenzyl)-3-carbamoylpyridinium chloride) and the reduction of a central ferric-iron of ferriprotoporphyrin IX by the above-described copolymer was studied spectrophotometrically in dimethyl sulfoxide. The rate of the reduction by the copolymer was much faster than by N-benzyl-3-carbamoyl-1,4-dihydropyridine. This acceleration by the copolymer could be explained by the intramolecular reduction of ferriprotoporphyrin IX which was coordinated by the pyridine residue of the copolymer.

Notes: We have investigated the B to A conformation change of DNA double helices by a new method “soft-mode analysis.” We find theoretically that a mode does soften when the vibration normal modes are perturbed by increasing the electrostatic interaction between the unbalanced charges on atoms in the double helix. The same mode also softens for enhanced van der Waals interactions. The mode softening indicates the onset of conformation change. The enhancing of the electrostatic and van der Waals interaction mimic the effect of decreasing the polar nature of the solvent or water of hydration associated with the B conformation DNA. We discuss qualitatively the concept of soft modes and their relation to conformation change as well as their applicability to macromolecules. We discuss previous work in which the normal vibrational modes have been calculated. We also discuss the displacement which comes from the soft mode and show that it correlates very well with that expected for the B to A conformation charge.

Notes: An electron microscope study of collagen fibrils from fixed tail tendons of rats has revealed that from some time shortly after birth until maturity, the fibril diameters have a bimodal distribution. The “two” types of fibril are indistinguishable in both transverse and longitudinal section. Unfixed specimens of eight-week-old-tail tendon showed a similar bimodal distribution of diameters though the positions of the peak values compared to fixed specimens of an eight-week-old-tail tendon were shifted upwards by about 30%. It has also been shown quantitatively that the polar collagen fibrils are directed randomly “up” and “down” with respect to their neighbors. Whilst it has been suggested by others that anastomosis is a feature of collagen structure, the results presented here do not support this hypothesis. Fibrillar units ∼ 140 Å in diameter have been observed and the possibilities that these are elastic fibers or the breakdown products of collagen fibrils have been considered.

Notes: The addition of a lytic layer to a preformed linear sucrose gradient induces a temporary (up to half a day) initial gradient of considerable steepness which retards the sedimentation of large (〉T4) DNAs. Centrifugation at sufficiently slow angular velocities permits the temporary initial gradient to disappear and therefore the sedimentation distance increases, yielding a rotor speed dependent effect on sedimentation distance. Gradients which are free of this effect are described and shown to permit mouse leukemia cell DNA to sediment independently of rotor speed (5-30 krpm).

Notes: The thermally induced helix-coil transitions of three A-T DNAs, d(A)n·d(T)n, d(A-T)n·d(A-T)n, and d(A-A-T)n·d(A-T-T)n, were studied. Experimental transition curves of the DNAs were analyzed using the loop entropy model of DNA melting. The calculation of the melting curve of d(A-A-T)n·d(A-T-T)n is presented using the integral equation formalism of Goel and Montroll. The aim of this work was to evaluate thermodynamic parameters which govern DNA stability and to test the theoretical model employed in the analysis. Our results show (1) an excellent over-all agreement between theory and experiment, (2) a loop entropy exponent k = 1.55 ± 0.05 provided the best fit to all the polymer transition curves, (3) the evaluated stacking free energies reflect the relative stability of the DNAs, and (4) the stacking energies of the ApA·TpT dimer evaluated from d(A)n·d(T)n and d(A-A-T)n·d(A-T-T)n differ. The last result is consistent with different conformations for the dimer in these two polymers.

Notes: Concentration and temperature dependences of the 1H nmr spectra of N-acetyl-L-proline N-methylamide were observed in various solvents [CCl4, CDCl3, (CD3)2CO, (CD3)2SO, H2O, and D2O]. The fraction of the cis isomer (with respect to the bond between the acetyl carbonyl carbon and prolyl nitrogen atoms) depends greatly on the solvent used; the fraction of the cis isomer is higher in polar solvents than in nonpolar solvents. It depends also on concentration and temperature in nonpolar solvents but not in polar solvents. In nonpolar solvents the trans isomer mostly exists in the γ-turn structure with an intramolecular hydrogen bond and the cis isomer tends to form molecular aggregates by intermolecular hydrogen bonds. In polar solvents both the cis and trans isomers exist in monomeric forms which interact with solvent molecules. The pH dependences of the N-methyl proton resonances indicate that the γ-turn structure of the trans isomer is present also in aqueous solution, though its population is difficult to determine. Apparent enthalpy and entropy changes for the conversion of the trans isomer to cis isomer are evaluated for various solvents. The results are discussed in terms of the intra- and intermolecular hydrogen bondings.

Notes: Cyclo(D-Leu-L-Leu) and cyclo(L-Leu-L-Leu) were synthesized, and their carbon-13 nmr spectra at 65 MHz were examined in dimethylsulfoxide and trifluoroacetic acid solutions. The chemical shift data are consistent with a boat or “twisted” boat conformation of the diketopiperazine ring in both solvents. There was no indication of protonation of the cyclic dipeptides by trifluoroacetic acid. Attempts at polymerizing the cyclic dipeptides were unsuccessful.

Notes: This paper presents new applications of the McMillan-Mayer solution theory to dispersions of highly charged colloidal cylinders in monovalent salt solutions. The thermodynamic solution properties are given in terms of the virial expansions relating to a Donnan membrane equilibrium. General expressions are derived for the second Donnan pressure virial coefficient B2 and for the first two salt distribution coefficients A1 and A2. The effect of electric interactions is represented as an increased effective diameter dB or dA of the colloidal cylinder. This yields the simple excluded volume expressions B2 = πdBL2/4 and A1 = πdA2L/4 for hard cylinders of length L and diameter dB and dA, respectively. The coefficient A2 is derived from the dependence of B2 on the salt concentration.Computations are made for double-stranded DNA in sodium chloride solutions with the DNA model developed in the preceding paper: a uniformly charged cylinder, with size and charge consistent with transport experiments, and surrounded by a Gouy double layer. In 1-0.005M sodium chloride solutions dB is found to vary from 29 Å to about 220 Å, and dA from 30 Å to about 170 Å, with little sensitivity to the uncertainties in the kinetic diameter d ≈ 24 Å and the experimental ζ potentials of DNA. Corresponding results predicted by the classical Donnan theory are 6-167 times too high for B2.Values of A2 are relatively small, in line with the expected rapid convergence of the virial expansion for the salt distribution. This is consistent with a phase transition from random to parallel orientation of the cylinders predicted first by Onsager for hard cylinders on the basis of B2, but not yet observed for DNA in simple salt solutions.

Notes: The interaction of ox ligamentum nuchae elastin (native, purified, and soluble) with different solvents (water, ethylene glycol, methanol, and 2,2,2-trifluoroethanol) was investigated by means of differential scanning calorimetry. The unfreezable solvent content and the corresponding solvent to elastin residue molar ratio were determined from the melting endotherms of the freezable solvent present in these systems. The molar ratio obtained in the case of water and ethylene glycol agrees with hypothesis of a direct solvation of the main chain peptide group and is interpreted according to a model previously proposed for polyamides. Quite different molar ratios are obtained in the case of the two monofunctional solvents and no model can be proposed at present to explain their interaction with the protein.

Notes: The synthesis and characterization of water-soluble random copolymers containing L-asparagine with N5-(4-hydroxybutyl)-L-glutamine, and the thermally induced helix-coil transitions of these copolymers in water, are described. The incorporation of L-asparagine was found to decrease the helix content of the polymers in water at all temperatures. The Zimm-Bragg parameters σ and s for the helix-coil transition in poly(L-asparagine) in water were deduced from an analysis of the copolymer melting curves in the manner described in earlier papers. The computed values of s indicate that asparagine destabilizes helical sequences at all temperatures in the range 0-60°C.

Notes: High-molecular-weight poly(0,0′-dicarbobenzoxy-L-β-3,4-dihydroxyphenyl-α-alanine) was prepared by the N-carboxyanhydride method. From the results obtained by a study of the optical rotation, nuclear magnetic resonance, and solution infrared absorption, the conformation of poly(0,0′-dicarbobenzoxy-L-β-3,4-dihydroxyphenyl-α-alanine) depended greatly on the solvent taking a right-handed helix with [θ]225 = -13,600 ∼ -18,900 in alkyl halides, a left-handed helix with [θ]228 = 22,100 ∼ 24,800 in cyclic ethers or trimethylphosphate, and a random coil structure in dichloroacetic acid, trifluoroacetic acid, or hexafluoroacetone sesquihydrate. The polypeptide underwent a right-handed helix-coil transition in chloroform/dichloroacetic acid (or trifluoroacetic acid) mixed solvents and a left-handed helix-coil transition in dioxane/dichloroacetic acid (or trifluoroacetic acid) mixed solvents. The results were compared with those of poly(0-carbobenzoxy-L-tyrosine).

Notes: Acetyl-(dehydro-Phe) and acetyl-bis(dehydro-Phe) groups have been attached to the ε-amino group of the lysine residues of the copolymer poly(Glu92Lys8) by reacting this last with acetyl-(dehydro-Phe)-azlactone and acetyl-bis(dehydro-Phe)-azlactone, respectively.In the latter case induced CD is observed between 250 and 330 nm, due to the relative dissymmetric disposition of the two dehydro-Phe groups under the chiral field of the polypeptide chain.pH dependence of the induced CD, observed for the copolymer and lacking in the lowmolecular-weight structural model, is related to the α-helical and random coiled conformation of the polypeptide chain.

Notes: A statistical theory of the linear dichroism of DNA-like chains is presented for two models which are discrete versions of the wormlike coil. In the final form the linear dichroism of the entire chain is related directly to the dichroic properties of a chain segment (base pair). Though the derivations are somewhat complicated, the result [Eq. (28)] is simple and the required statistical parameters can be easily calculated for either model from measured values of the persistence length. In fact, for molecules as stiff as double-stranded DNA, the results can be reduced with good accuracy to the form \documentclass{article}\pagestyle{empty}\begin{document}$$ \frac{{\Delta \varepsilon (chain)}}{{\Delta \varepsilon (segment)}} = \frac{{P_{\infty} }}{l}\frac{r^{2}}{{\langle r^2 \rangle_0 }} $$\end{document} showing that the “optical persistence” given on the left is directly proportional to the structural persistence, P∞/l. As in previous theories the results are restricted to chains in their Gaussian limit.

Notes: A histidine-containing cyclic dipeptide, cyclo(D-Leu-L-His), was almost 20 times as efficient a catalyst as imidazole in the hydrolysis of p-nitrophenyl laurate. The effect of dioxane on the hydrolysis showed that hydrophobic interaction between the cyclic dipeptide and the ester is very important. This reaction obeyed the Michaelis-Menten kinetics, and the Michaelis constant Km was as low as 9.98 × 10-5M. Since the linear dipeptide having D-Leu-L-His sequence was nearly inactive in the hydrolysis, the functional groups of cyclo(D-Leu-L-His) in a specific arrangement held by the rigid backbone must have cooperated in the fast hydrolysis. Very weak catalysis by the diasteremeric cyclic dipeptide, cyclo(L-Leu-L-His), in the hydrolysis supported the above view.

Notes: The charge-transfer complexes of a poly(L-tryptophan) sequence with imidazolium hydrochloride and poly(L-histidine hydrochloride) have been investigated in 2,2,2-trifluoroethanol by ultraviolet (uv), circular dichroism (CD), and fluorescence techniques. Both complexes exhibit absorption maxima centered at around 275 nm, whereas hypochromism with respect to the combined spectra of the constituents can be observed below 250 nm. All complexes show optical activity in the near uv and in the peptide absorption region, which is discussed in terms of the conformational properties of the donor. A marked decrease of the fluorescence intensity of the L-tryptophan sequence is observed upon addition of imidazolium hydrochloride and poly(L-histidine hydrochloride). From the fluorescence data the formation constant of the charge-transfer complex between the L-tryptophan sequence and imidazolium ions is also evaluated.

Notes: Spectrophotometric titrations of slightly substituted carboxymethylamylose (CM-Amy) and diethylaminoethylamylose (DEAE-Amy) with iodine in the presence of iodide (I2/I-) were carried out as a function of iodide concentration, temperature, and polymeric charge. Binding isotherms for the polymer-I2/I- complex are reported in terms of an apparent binding constant (Ka) plotted versus degree of saturation of the complex (θ). The dependence of Ka upon polymeric charge is interpreted as evidence for the negatively charged character of the bound species. The cooperative nature of the binding process is evident in the positive slope of Ka vs (θ). Whereas the apparent binding constants and binding cooperativities for the derivatives are smaller than for the amylose-I2/I- complex, the binding enthalpies deduced from the temperature dependence of Ka at θ = 0.5 appear to be the same for amylose and CM-Amy. A viscometric titration of fully charged CM-Amy with I2/I-, conducted at dialysis equilibrium between the CM-Amy-I2/I- solution and the polymer-free solvent phase, disclosed a maximum in the plot of intrinsic viscosity ([η]) vs θ. The increase in [η] at small θ was interpreted as a reflection of polyelectrolyte expansion provoked by absorption of the negatively charged bound species; the subsequent decline in [η] is attributed to stabilization by I2/I- of compact helical sequences or to the formation at higher θ of intermolecular aggregates.

Notes: The effect of various salts on the enzymatic activity of beef-liver glutamate dehydrogenase, on the binary enzyme-reduced coenzyme (NADH or NADPH) comples, as well as on the ternary complex with glutamate was investigated in aqueous solution (0.067M phosphate buffer, pH 7.6). Binding studies in the analytical ultracentrifuge and circular dichroism measurements indicated dissociation of the coenzyme from the enzyme-coenzyme complex by the action of various salts. The efficiency of this change was largely dependent on the type of anion present and generally followed the series: acetate 〈 Br- 〈 I- 〈 SCN-. Acetate ions and guanidinium thiocyanate showed exceptional behaviour in some cases, while K+ and Na+ gave similar results. The reversibility of the ion effects on the enzymatic activity was demonstrated by dilution tests. Upon addition of salts, the inhibitory effect of GTP was slightly changed in most cases, while the activating effects of ADP and L-leucine were practically abolished and with ADP, the halides caused an additional inhibition. Assuming an equilibrium involving dissociation of the enzyme-coenzyme complex by the action of salts, an exponent of 1.3 for NaBr and of 2.0 for KSCN was calculated for the respective concentrations. The apparent equilibrium constants were evaluated to be about 20 times greater for KSCN than for NaBr.

Notes: Poly(Glu(OBzl)-Gly)n, poly(Glu-Gly)n, poly(Gly)-(Glu(OBzl)-Gly), and poly(Gly-Glu-Gly) were synthesized from the pentachlorophenyl esters of the sequential monomer. Both of the polymers containing free glumatic-acid residues are soluble in water, as is the lower molecular weight fraction of the polytripeptides with the benzyl ester in place. Circular dichroism studies and infrared dichroism studies suggest that the 21 helix is favored for the polydipeptide with removal of the benzyl ester reducing the conformational integrity. The polytripeptide showed evidence of 31 helix in addition to the 21 form, depending on solvent. A rationale for the conformations observed is developed based on the bulkiness of the side-chain residues and conformational stabilization, in certain cases, by hydrophobic interactions between the benzyl ester groups.

Notes: The thermoelastic behavior of water solvated elastin has been investigated in simple tension, in the temperature range 0-70°C. Specimens purified from both the ox ligamentum nuchae and pig thoracic aorta have been studied. Force data obtained by cycling the temperature for various constant specimen lengths display a separated variable dependence of the form f = A(T)B(α), where T is absolute temperature and α the extension ratio. For ligament elastin B(α) is a linear function whereas for aortic elastin it is a nonlinear function. The applicability of the rubber elasticity theory to elastin has been tested by setting A(T) equal to the temperature-dependent front factor for simple tension of a homogeneous rubber whilst B(α) is left undefined. In this way it has been possible to take into account the fibrous nonhomogeneity of the polymer, and also to avoid any inconsistency within the theory of attributing a dependence of the variable fe/f upon extension ratio. The behavior of both ligament and aortic elastin agrees well with the conclusion that the dominant deformation mechanism is entropy elastic, fe/f ≪ 1. The linearity of the load isotherm for ligament elastin permits a particularly simple experimental procedure using a single force-temperature plot for one value of interclamp length. Using this procedure high precision has been obtainble, and the data shows a close adherence to the theory with fe/f = 0.1. The relationship between this result and current controversy over the molecular conformation of elastin is discussed.

Notes: Methods are given for analyzing regularly spaced patterns of amino acids in proteins and applied to the α1 chain of collagen. Fourier methods use the transform of the sequence either embedded in a very long array or folded onto a fundamental base period. Filtering through a moveable “window” of definite width is used to display almost regular features at any chosen frequency. A pattern detection method is described for patterns of general shape. Collagen has statistically significant periodicities at fractions of the stagger distance D = 670 Å. Hydrophobic groups show strong orders of 5, 6, 11; proline 5; charged groups 6, 18, 21. Charged residues mostly occur as neutral pairs. Their distribution has strong 6th and 21st orders which also appear in the changes which are paired at multiples of D. Charge pairs separated by (D + 3) residues show a strong 5D/89 pattern and may form a system of salt bridges across the fibril. There is no sign of any regular pattern of amino acids over the triple helix with a period close to its natural pitch of 30 residues. Supercoiled models with six relative turns of the contact edge between paired triple-helical strands are examined.

Notes: Circular dichroism spectra have been obtained for cationic poly(L-arginine) and poly(L-histidine) in aqueous solutions containing varying amounts of sodium dodecyl sulfate. The detergent induces a disorder-order transition in both polypeptides. In each case the transition is cooperative and occurs when the ratio of detergent to amino acid residue is near unity. The ordered structure formed by poly(L-arginine) is readily identifiable as an α helix. Poly(L-histidine) appears to form a β structure in which the 211-nm electronic absorption band of the imidazole group exhibits significant rotatory strength.

Notes: We have measured the circular dichroism (CD) and absorption properties of poly[r(G-U)] and poly [d(G-T)] over a wide range of Na+ concentrations and temperatures. We find evidence for self-complexed forms of these polymers at lower temperatures and/or higher Na+ concentrations than generally needed for double-strand formation in other DNA and RNA polymers. These self-complexes could be composed of double-stranded regions with weak G·U or G·T base pairs.

Notes: Comparative studies on the interaction of 8-anilino-1-naphthalenesulfonate (ANS) with polylysine and polyarginine have been made by equilibrium dialysis and fluorescence or circular dichroism measurements, to investigate the structural characteristics of the polypeptides. The results are summarized as follows: (i) ANS binds to either of the polypeptides primarily by electrostatic interaction while hydrophobic interaction partially facilitates the dye binding; both interactions are stronger in the polyarginine-dye binding than the polylysine-dye binding. (ii) The fluorescence of ANS is more intensified when the dye binds to polyarginine than to polylysine regardless of the value of r (number of bound dye per amino-acid residue) of polypeptide-dye complexes, although the intensification depends on the r value and becomes maximum at r = 0.25-0.35 for both cases. (iii) The binding of ANS to each polypeptide is cooperative at r 〈 0.4. (iv) The circular dichroism is more efficiently induced in the spectral region of ANS by binding to polyarginine than to polylysine.From these results, it was concluded that, compared to polylysine, polyarginine suffers some structural change by ANS binding into a more compact molecular configuration having some regularity with a lower dielectric environment.

Notes: Circular dichroism (CD) spectra of poly(dA), poly(dT), poly(dA)·poly(dT), and poly[d(A-T)]·poly[d(T-A)] have been measured as a function of temperature. From these data difference spectra have been calculated by subtracting the spectrum measured at low temperature from the spectra measured at higher temperatures. The CD difference spectra obtained upon melting of the two double-stranded polymers are very similar. From a comparison of these difference spectra with calculated ones it is shown that optical transitions near 272 nm (on A) and 288 nm (most probably on T) are present. The premelting changes of the CD spectrum of poly[d(A-t)]·poly[d(T-A)] are due to a change in conformation in which the secondary structure goes from a C- to B-type spectrum by increasing the A-type nature of the polymer. Such a change is not observed for poly(dA)·poly(dT). Instead, a transition between two different B-type geometries occurs.

Notes: The solution characterization of poly(Lys-Ala-Glu) is described. This polytripeptide is zwitterionic at neutral pH and is shown to take on a conformation which is dictated by the state of ionization, molecular weight, temperature, and solvent. The polypeptide is almost entirely α-helical at low pH and temperature for polymers of greater than 25,000 molecular weight. Melting profiles for these conditions show tm ∼ 20°C. Analysis of circular dichroism curves shows the α-helical content to vary in a linear manner with molecular weight in the range 3000-30,000. At neutral pH the charged polypeptide is essentially random, but substantial α-helix could be induced by addition of methanol or trifluoroethanol. At temperatures where the sequential polypeptide is a random coil, addition of trifluoroethanol produces a polymer which is mostly α-helical but also contains an appreciable ammount of β-structure. The infrared spectrum of a low-molecular-weight fraction assumed to be cyclo(Lys-Ala-Glu)2 was tentatively assigned a β-pleated sheet structure.A comparison of this polytripeptide in various ionization states with other polytripeptides containing L-alanine and L-glutamate or L-lysine shows the α-helix directing properties for the (uncharged) residues to lie in the order Ala 〉 Glu 〉 Lys.

Notes: Previously used procedures for processing the amino acids from 6N hydrochloric acid hydrolysis of poly[N5-(4-hydroxybutyl)-L-glutamine], poly[N5-(3-hydroxypropyl)-L-glutamine], and several random copolymers derived from these, led to the formation of spurious products. These have been isolated and characterized as the γ-ester of glutamic acid and the hydroxyalkyl amine, and chloro-alkyl amine hydrochloride. The former reduces the observed values for glutamic acid, but the latter has no effect on them. A method is used to avoid formation of these artifacts in the amino-acid analysis. Of all the copolymers studied previously in this series, the compositions of only those containing L-serine are in error as a result of the formation of the γ-ester. A redetermination of the amino-acid compositions of the copolymer fractions studied earlier leads to slightly revised values for the Zimm-Bragg parameters σ and s of serine.

Notes: A new technique is presented for treating the ground state of an heteropolymer with a random sequence of components. An exact system of equations is found for determining the ground state energy E which is equal to the polymer free energy f in the lowest-order approximation in T/V (V/2 is the large “surface” energy arising at the boundaries between coiled and “helical” sections: V ≫ T, Uk; U1 and -U2 are the free energies of the components counted from the corresponding coiled state energies). These equations are essentially simplified at certain fixed values of the ratio U1/U2. For integer values of U2/U1 and U1/U2 a solution is obtained with an accuracy exp(-V/Uk). The ground-state energy as a function of U1 and U2 is shown to be highly irregular: its derivatives have jumps at an infinite number of points. These jumps provide a fine structure of the melting curves. A smoothed over the jumps function E′ is found by way of analytic continuation from the integer values of U1/U2 and U2/U1. The accuracy of the approximation f ≈ E is estimated and the correctional term of order T/V is determined.

Notes: In order to obtain information about the conformational characteristics at the nearestneighbor level in the 2′-O-methylated region of t-RNA, as well as in the bizarre 5′-terminus of eucaryotic mRNA, a detailed nuclear magnetic resonance study of 2′-O-methyl-cytidylyl-(3′ → 5′)-cytidine (CmpC) was conducted. Proton spectra were recorded at 270 MHz in the Fourier mode in D2O solutions, 0.01M, pD 7.3 in the temperature range 5-80°C. Complete accurate sets of nmr parameters were derived for each of the nucleotidyl units by a combination of homo-nuclear decouplings and simulation iteration methods. The data were translated into conformational parameters using procedures developed in earlier studies from these laboratories.It is shown that the ribofuranose ring exists at a 2E ⇄ 3E equilibrium with clear preference [(75-80)%] for the 3E mode. The C(4′)-C(5′) and C(5′)-O(5′) bonds form a stable conformational network with outspoken preference for conformers in which Ψ1, Ψ2 ≃ 60° and φ2 ≃ 180°. The orientation of the 3′-phosphate and 2′-O-methyl groups is such that φ1′ ≃ 210° and φ″ ≃ 60°. The phosphodiester bonds are flexible and shift trends for base, H(1′), and H(5″) suggest the existence of a conformational blend of right-handed stack (g-g-), left-handed stack (g+g+), and unstacked arrays (tg- and tg+).Elevation of temperature perturbs the 2E ⇄ 3E equilibrium accompanied with modest depopulation of ψ1, ψ2 ≃ 60° and φ2 ≃ 180° conformers. The major effect of elevation of temperature is in the increase of unstacked arrays at the expense of g-g- and g+g+ conformers. The shift trend of Cmp-H(3′) with temperature shows that torsional variation about O(3′)-P is facilitated by increase in temperature and the preferred rotamer about O(3′)-P in the unstacked form is t (ω1′ = 180°).A detailed comparison of the aqueous solution conformations of CpC and CmpC reveals that 2′-O-methylation causes: (i) a reduction in the magnitude of χ1; (ii) an increase in the population of 3E pucker at the 3′-nucleotidyl unit; and (iii) modest perturbations in the O(3′)-P and P-O(5′) bond conformations. Comparison of the aqueous solution conformations of AmpA and CmpC makes clear that the conformational properties of pyrimidine-pyrimidine and purine-purine dimers which carry a 2′-O-methylated 3′-nucleotidyl unit are significantly different.

Notes: Poly(Lys(Cbz)-Ala-Glu(OBzl)) was prepared by the self-condensation of Lys(Cbz)-Ala-Glu(OBzl)-ONSu in dimethylformamide. After deprotection of the side chains, the product was subjected to Sephadex G-50 chromatography. The molecular weight of unfractionated and fractionated poly(Lys-Ala-Glu) was calculated from a calibrated Sephadex G-50 column, spectrophotometrically from Dnp-(Lys-Ala-Glu), equilibrium centrifugation, and viscosity measurements. Approximately 21% of the unfractionated material was polymeric with the remaining 79% being cyclic and monomeric material. Treatment of polymer hydrolysate with L-amino acid and D-amino acid oxidase indicated poly(Lys-Ala-Glu) to be optically pure. The apparent pKa's of the two ionizable groups were 4.1 and 9.7.

Notes: Yeast tRNA3Leu is one of several tRNA molecules which can adopt a stable, biologically inactive, denatured conformation. The circular dichroism of the native and denatured conformers differs, providing the basis for the present study of the mechanism for the renaturation process. Conversion of the denatured structure to the native takes place in two steps: a rapid change occurring immediately on addition of Mg++, followed by a slower, strongly temperature-dependent step which returns the molecule to its biologically active state. Optimal kinetic data for the second step could be obtained at 285 nm. Analysis of the time dependence of Δε285 by the Guggenheim method demonstrated that this step follows first-order kinetics. The temperature dependence of the rate constants over the range 32-41°C yielded the following parameters for the rate-limiting step: Ea = 69 kcal/mole, ΔH

Notes: The methods suggested earlier for the analysis and representation of protein structural data are now extended to the helical regions in finer details. These enable better handling of characterization of bends and distortions, for which statistical parameters are also developed. Using latest myoglobin data, best experimental parameters for the α-helix are deduced to be rN = 1.55 (0.13) Å, rCα = 2.28 (0.12) Å, rC′ = 1.70 (0.10) Å, r0 = 2.02 (0.12) Å, φ = 100.5 (2.3)°, and t = 1.495 (0.055) Å.

Notes: We have monitored the helix-coil transition of the self-complementary d-CpCpGpG and d-GpGpCpC sequences (20mM strand concentration) at the base pairs, sugar rings, and backbone phosphates by 360-MHz proton and 145.7-MHz phosphorus nmr spectroscopy in 0.1M phosphate solution between 5 and 95°C. The guanine 1-imino Watson-Crick hydrogen-bonded protons, characteristic of the duplex state, are observed below 10°C, with solvent exchange occurring by transient opening of the tetranucleotide duplexes. The cytosine 4-amino Watson-Crick hydrogen-bonded protons resonate 1.5 ppm downfield from the exposed protons at the same position in the tetranucleotide duplexes, with slow exchange indicative of restricted rotation about the C-N bond below 15°C. The guanine 2-amino exchangeable protons in the tetranucleotide sequence exhibit very broad resonances at low temperatures and narrow average resonances above 20°C, corresponding to intermediate and fast rotation about the C-N bond, respectively. Solvent exchange is slower at the amino protons compared to the imino protons since the latter broaden out above 10°C. The well-resolved nonexchangeable base proton chemical shifts exhibit helix-coil transition midpoints between 37 and 42°C. The transition midpoints and the temperature dependence of the chemical shifts at low temperatures were utilized to differentiate between resonances located at the terminal and internal base pairs while the H-5 and H-6 doublets of individual cytosines were related by spin decoupling studies. For each tetranucleotide duplex, the cytosine H-5 resonances exhibit the largest chemical shift change associated with the helix-coil transition, a result predicted from calculations based on nearest-neighbor atomic diamagnetic anisotropy and ring current contributions for a B-DNA duplex. There is reasonable agreement between experimental and calculated chemical shift changes for the helix-coil transition at the internal base pairs but the experimental shifts exceed the calculated values at the terminal base pairs due to end-to-end aggregation at low temperatures. Since the guanine H-8 resonances of the CpCpGpG and d-CpCpGpG sequences exhibit upfield shifts of 0.6-0.8 and 〈0.1 ppm, respectively, on duplex formation, these RNA and DNA tetranucleotides with the same sequence must adopt different base-pair overlap geometries. The large chemical shift changes associated with duplex formation at the sugar H-1′ triplets are not detected at the other sugar protons and emphasize the contribution of the attached base at the 1′ position. The coupling sum between the H-1′ and the H-2′ and H-2″ protons equals 15-17 Hz at all four sugar rings for the d-CpCpGpG and d-GpGpCpC duplexes (25°C), consistent with a C-3′ exo sugar ring pucker for the deoxytetranucleotides in solution. The temperature dependent phosphate chemical shifts monitor changes in the ω,ω′ angles about the O-P backbone bonds, in contrast to the base-pair proton chemical shifts, which monitor stacking interactions.

Notes: A strong, positive, extrinsic CD band ([θ]242.5 = ∼2 × 10-3 deg cm2/dmole) has been observed for a α-bromo-poly[methylene-1,4-phenylenecarbonyloxyethylene(dimethylamino) bromide] (I). The extrinsic Cotton effect is attributed to the ordered arrangement of the aromatic chromophores along the DNA helix.The extrinsic band had a linear dependence on the amount of polycation I added from r ≤ 0.3 to r = ∼0.5, but decreased thereafter. Addition of the polycation decreased the positive CD band of DNA at 275 nm. The transformation of B → C form in the presence of salts or other polycations caused similar changes. The decrease in [θ]275 was reversed at higher concentrations of the polycation (r 〉 0.4).Thermal denaturation studies indicated both stabilization of the helix conformation (Δtm = 21°C) and a high degree of cooperativity in the melting of DNA-polycation complex as compared to native calf thymus DNA. Using the linear relationship between r (polycation residue/DNA phosphate) and F (fraction of bound base pairs), a value of 0.6 was derived for β (number of monomer residues of polycation/nucleotide).Both electrostatic and hydrophobic effects probably influence the stability of the DNA-polycation complex, since the strength of the 242.5 nm CD band is a function of both salt and urea concentrations.

Notes: From the results of 13C-nmr measurement of poly(β-benzyl-L-aspartate) and its model compounds in dimethyl sulphoxide/deuterated chloroform mixtures, it was found that the side chain of poly(β-benzyl-L-aspartate) is solvated by dimethyl sulphoxide in the region more than dimethyl sulphoxide 20% (v/v), where the backbone maintains the α-helix. The chemical shift differences in the benzyl group carbons of poly(γ-benzyl-L-glutamate) (trifluoroacetic acid/deuterated chloroform) accompanied by the helix-coil transition, originate from the interaction between the ester group of the side chain and trifluoroacetic acid. The chemical shift difference in the ester carbon is similar. On the other hand, the chemical shift differences of the side-chain carbons in the alkyl portion (Cβ, Cγ) originate not only from the interaction between the ester group of the side chain and trifluoroacetic acid, but also from some other unknown factors. The chemical shift differences of the side-chain carbons of poly(β-benzyl-L-aspartate) originate from the interaction between the ester group of the side chain and trifluoroacetic acid.

Notes: The dielectric properties of DNA solutions at low frequencies (5 Hz to 2 kHz) have been measured by means of a four-terminal bridge method utilized to minimize electrode polarization errors. At 24°C native salt-free DNA has a very large specific dielectric increment, Δε/c = 9.8 × 106 l/mol and a very low frequency relaxation centered at 18 Hz. Both the dielectric increment and the relaxation time are greatly decreased by partial heat denaturation at temperatures above 60°C or by addition of salt, the effects being much larger for divalent anions. These results are shown to be in qualitative agreement with theoretical treatments of counterion fluctuation polarization by McTague and Gibbs for the equilibrium case and by Mandel for relaxation. The ratio of the relaxation time for the low-frequency process to that previously observed at much higher frequencies suggests that these relaxations result from counterion fluctuations along the longitudinal and transverse axes of the molecule, respectively.