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  • 1
    Electronic Resource
    Electronic Resource
    Developmental regulatory elements in the 5′ flanking DNA of the Drosophila choline acetyltransferase gene (1993)
    Springer
    Development genes and evolution 202 (1993), S. 159-169 
    Details
    ISSN: 1432-041X
    Keywords: Drosophila ; Choline acetyltransferase ; cis-Regulatory element ; lacZ reporter gene ; Colinergic neuron
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Choline acetyltransferase (ChAT, EC 2.3.1.6) catalyzes the production of the neurotransmitter acetylcholine, and is an essential factor for neurons to be cholinergic. We have analyzed regulation of the Drosophila ChAT gene during development by examining the β-galactosidase expression pattern in transformed lines carrying different lengths of 5′ flanking DNA fused to a lacZ reporter gene. The largest fragment tested, 7.4 kb, resulted in the most extensive expression pattern in embryonic and larval nervous system and likely reflects all the cis-regulatory elements necessary for ChAT expression. We also found that 5′ flanking DNA located between 3.3 kb and 1.2 kb is essential for the reporter gene expression in most of the segmentally arranged embryonic sensory neurons as well as other distinct cells in the CNS. The existence of negative regulatory elements was suggested by the observation that differentiating photoreceptor cells in eye imaginal discs showed the reporter gene expression in several 1.2 kb and 3.3 kb transformants but not in 7.4 kb transformants. Furthermore, we have fused the 5′ flanking DNA fragments to a wild type ChAT cDNA and used these constructs to transform Drosophila with a Cha mutant background. Surprisingly, even though different amounts of 5′ flanking DNA resulted in different spatial expression patterns, all of the positively expressing cDNA transformed lines were rescued from lethality. Our results suggest that developmental expression of the ChAT gene is regulated both positively and negatively by the combined action of several elements located in the 7.4 kb upstream region, and that the more distal 5′ flanking DNA is not necessary for embryonic survival and development to adult flies.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00365306
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  • 2
    Electronic Resource
    Electronic Resource
    Localization of choline acetyltransferase-expresing neurons in the larval vissual system of Drosophila melanogaster (1995)
    Springer
    Cell & tissue research 282 (1995), S. 193-202 
    Details
    ISSN: 1432-0878
    Keywords: Choline acetyltransferase ; Cholinergic neuron ; Visual system ; Bolwig's organ ; Immunocytochemistry ; In situ hybridization ; Drosophila melanogaster (Insecta)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Choline acetyltransferease (ChAT) is the enzyme catalyzing the biosynthesis of acetylcholine and is considered to be a phenotypically specific marker for cholinergic neurons. We have examined the distribution of ChAT-expressing neurons in the larval nervous system of Drosophila melanogaster by three different but complementary techniques: in situ hybridization with a cRNA probe to ChAT messenger RNA, immunocytochemistry using a monoclonal anti-ChAT antibody, and X-gal staining of transformed animals carrying a reporter gene composed of 7.4 kb of 5′ flanking DNA from the ChAT gene fused to a lacZ reporter gene. All three techniques demonstrated ChAT-expressing neurons in the larval visual system. In embryos, the photoreceptor organ (Bolwig's organ) exhibited strong cRNA hybridization signals. The optic lobe of late third-instar larvae displayed ChAT immunoreactivity in Bolwig's nerve and a neuron close to the insertion site of the optic stalk. This neuron's axon ran in parallel with Bolwig's nerve to the larval optic neuropil. This neuron is likely to be a first-order interneuron of the larval visual system. Expression of the lacZ reporter gene was also detected in Bolwig's organ and the neuron stained by anti-ChAT antibody. Our observations indicate that acetylcholine may be a neurotransmitter in the larval photoreceptor cells as well as in a first-order interneuron in the larval visual system of Drosophila melanogaster.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00319111
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  • 3
    Electronic Resource
    Electronic Resource
    Developmental expression of choline acetyltransferase mRNA inDrosophila (1990)
    Springer
    Neurochemical research 15 (1990), S. 1089-1096 
    Details
    ISSN: 1573-6903
    Keywords: Choline acetyltransferase ; development ; mRNA ; Drosophila
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract We have measured the steady state levels of choline acetyltransferase (ChAT, EC 2.3.1.6) mRNA during different developmental stages ofDrosophila melanogaster using a ChAT specific cRNA probe. ChAT mRNA was first detected approximately 6–7 h after oviposition, increased until the 1st–2nd larval instar, decreased into early pupal stages and increased again during late pupation, reaching a maximum in adults. Northern analysis showed a major RNA band with a Mr of 4.7 kilobases and Western analysis also showed a single major 75 kD protein band at all developmental stages. Our results support the hypothesis that a major point of regulation of ChAT expression may be at the transcriptional level.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF01101709
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  • 4
    Electronic Resource
    Electronic Resource
    Drosophila Choline Acetyltransferase Temperature-Sensitive Mutants (1999)
    Springer
    Neurochemical research 24 (1999), S. 1081-1087 
    Details
    ISSN: 1573-6903
    Keywords: Choline acetyltransferase ; Drosophila ; Temperature-sensitive mutants
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract We used the reverse transcription-polymerase chain reaction (RT-PCR) to amplify choline acetyltransferase (ChAT) mRNA fragments from two temperature-sensitive alleles of Drosophila melanogaster, Cha ts1 and Cha ts2. Single base substitutions in the mutants (T1614A in Cha ts1 and G1596A in Cha ts2) would result in amino acid changes for ChAT protein (Met403Lys in Cha ts1 and Arg397His in Cha ts2). These base substitutions were confirmed in mRNA extracted from homozygous mutants using a Single Nucleotide Primer Extension assay (SNuPE) and are sufficient to produce thermolabile enzyme. Our results indicate that these temperature-sensitive mutants are point mutations in the structural gene for ChAT. Using a quantitative SNuPE assay we also show that similar levels of Cha ts and wild type transcripts are present in heterozygous flies (Cha ts1/+ and Cha ts2 /+) at both restrictive and permissive temperatures. This contrasts with RNase protection assays of ChAT mRNA in homozygous mutant animals where the levels of mutant mRNA decrease at restrictive temperature.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1021021213625
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  • 5
    Electronic Resource
    Electronic Resource
    Kinetics of ethidium's intercalation in packaged bacteriophage T7 DNA: Effects of DNA packing density (1986)
    New York : Wiley-Blackwell
    Biopolymers 25 (1986), S. 1345-1357 
    Details
    ISSN: 0006-3525
    Keywords: Chemistry ; Polymer and Materials Science
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology
    Notes: The kinetics of ethidium's intercalative binding to DNA packaged in bacteriophage T7 and two T7 deletion mutants have been determined, using enhancement of fluorescence to quantitate binding. At a constant ethidium concentration, the results can be described as first-order binding with two different rate constants, k1* (= k1 + k-1) and k2* (= k2 + k-2). The larger rate constant (k1*) was at least four orders of magnitude smaller than the comparable first-order forward rate constant for binding to DNA released from its capsid. At 25°C values of k1* decreased as the amount of DNA packaged per internal volume increased. This latter observation indicates that the rate of ethidium's binding to packaged T7 DNA is limited by an event that occurs inside of the DNA-containing region of T7, not by the crossing of T7 capsid's outer shell. Arrhenius plots of kM1* are biphasic, indicating a transition for packaged DNA at a temperature of 20°C. The data indicate that k1* s are limited by either sieving of ethidium during its passage through the packaged DNA or subsequent hindered intercalation.
    Additional Material: 7 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/bip.360250713
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  • 6
    Electronic Resource
    Electronic Resource
    The presence in collagen of γ-glutamyl peptide linkagesSupported by grants from the National Institutes of Health, U. S. Public Health Service, A-3172, H-4762, H-3838, M-2565, M-2562. The material in this paper is taken from a thesis submitted by Carl Franzblau in partial fulfillment of the requirements for the degree of Doctor of Philosophy in the Sue Golding Graduate Division of Medical Sciences, Albert Einstein College of Medicine, Yeshiva University. (1963)
    New York : Wiley-Blackwell
    Biopolymers 1 (1963), S. 79-97 
    Details
    ISSN: 0006-3525
    Keywords: Chemistry ; Polymer and Materials Science
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology
    Notes: A method is described for formation of hydroxamic acids by direct coupling, in aqueous medium, of hydroxylamine hydrochloride and the sodium salt of a carboxylic acid. The reaction is mediated by a water-soluble carbodiimide, 1-cyclohexyl-3-[2-morpholinyl-(4)-ethyl]-carbodiimide metho-p-toluene sulfonate. Using model compounds, the production of α-, β- and γ-acyl hydroxamic acids was studied. Hydroxamic acid derivatives of α-polyglutamic and γ-polyglutamic acids were also prepared. Dinitro-phenylhydroxamate derivatives were prepared and subjected to Lossen rearrangement, and the reaction mixtures hydrolyzed. Analysis showed that α-polyglutamic acid yielded as many molecules of α,γ-diaminobutyric acid as there had been glutamic acid residues in the form of the dinitrophenylhydroxamate, and gave rise to no ammonia and succinic semialdehyde. In contrast, γ-polyglutamic acid yielded exactly twice the molar quantity of ammonia as there had been glutamic acid residues in the form of dinitrophenylhydroxamate, and also gave rise to significant quantity of succinic semi-aldehyde but no α,γ-diaminobutyric acid. Since production of the latter is characteristic of α-glutamyl hydroxamates and production of the former is associated with γ-glutamyl hydroxamates, the results indicated that the side-chain carboxyl groups of either polymer retained their identities during formation of the hydroxamic acid derivative, and no interchange had occurred between α- and γ-carboxyl groups. These methods were then used to establish that gelatin of ichthyocol contains (per 1000 total residues) at least 20 residues of glutamic acid in γ-peptide linkage. Due to the incomplete dinitrophenylation of the hydroxamic acid derivative of the protein, gelatin of calf skin collagen gave a lower figure of 10 such linkages per 1000 total amino acid residues-a value which must be considered a minimum value. These results show that γ-glutamyl peptide bonds exist in collagen.
    Additional Material: 1 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/bip.360010109
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  • 7
    Electronic Resource
    Electronic Resource
    Polyphenylquinoxalines containing pendant phenylethynyl groups: Preliminary mechanical properties (1983)
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Applied Polymer Science 28 (1983), S. 355-366 
    Details
    ISSN: 0021-8995
    Keywords: Chemistry ; Polymer and Materials Science
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology , Mechanical Engineering, Materials Science, Production Engineering, Mining and Metallurgy, Traffic Engineering, Precision Mechanics , Physics
    Notes: Polyphenylquinoxalines (PPQ's) containing various amounts of crosslinkable pendant phenylethynyl groups were prepared from the reaction of 3,3′,4,4′-tetraaminodiphenyl ether, 4,4′-oxydibenzil, and 4,4′-oxybis(4″-phenylethynylbenzil). The distribution of the pendant phenylethynyl groups along the polymer chain was varied in an attempt to alter the properties of the polymers. Preliminary film, adhesive, and laminate properties of PPQ void of pendant crosslinkable groups and containing pendant phenylethynyl groups were determined. The thermally induced reaction of the phenylethynyl group crosslinked the polymer which resulted in better dimensional stability at elevated temperatures.
    Additional Material: 5 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/app.1983.070280131
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  • 8
    Electronic Resource
    Electronic Resource
    An analysis of side chain interactions and pair correlations within antiparallel β-sheets: The differences between backbone hydrogen-bonded and non-hydrogen-bonded residue pairs (1995)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 22 (1995), S. 119-131 
    Details
    ISSN: 0887-3585
    Keywords: antiparallel β-sheet ; twist ; protein folding ; side chain interactions ; branched amino acids ; cystine-rich proteins ; side chain packing ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Cross-strand pair correlations are calculated for residue pairs in antiparallel β-sheet for two cases: pairs whose backbone atoms are hydrogen bonded together (H-bonded site) and pairs which are not (non-H-bonded site). The statistics show that this distinction is important. When glycine is located on the edge of a sheet, it shows a 3:1 preference for the H-bonded site. Thestrongest observed correlations are for pairs of disulfide-bonded cystines, many of which adopt a close-packed conformation with each cystine in a spiral conformation of opposite chirality to its partner. It is likely that these pairs are a signature for the family of small, cystine-rich proteins. Most other strong positive and negative correlations involve charged and polar residues. It appears that electrostatic compatibility is the strongest factor affecting pair correlation. Significant correlations are observed for β- and γ-branched residues inthe non-H-bonded site. An examination of the structures showsa directionality in side chain packing. There is a correlation between (1) the directionality in the packing interactions of non-H-bonded β- and γ-branched residue pairs, (2) the handedness of the observed enantiomers of chiral β-branched side chains, and (3) the handedness of the twist of β-sheet. These findings have implications for the formation of β-sheets during protein folding and the mechanism by which the sheet becomes twisted. © 1995 Wiley-Liss, Inc.
    Additional Material: 7 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/prot.340220205
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  • 9
    Electronic Resource
    Electronic Resource
    Towards meeting the paracelsus challenge: The design, synthesis, and characterization of paracelsin-43, an α-helical protein with over 50% sequence identity to an all-β protein (1996)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 24 (1996), S. 502-513 
    Details
    ISSN: 0887-3585
    Keywords: de novo design ; protein structure ; inverse folding ; genetic algorithms ; 1H NMR ; CD ; peptide ; protein folding ; methanol ; ethylene glycol ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: In response to the Paracelsus Challenge (Rose and Creamer, Proteins, 19:1-3, 1994), we present here the design, synthesis, and characterization of a helical protein, whose sequence is 50% identical to that of an all-β protein. The new sequence was derived by applying an inverse protein folding approach, in which the sequence was optimized to “fit” the new helical structure, but constrained to retain 50% of the original amino acid residues. The program utilizes a genetic algorithm to optimize the sequence, together with empirical potentials of mean force to evaluate the sequence-structure compatibility. Although the designed sequence has little ordered (secondary) structure in water, circular dichroism and nuclear magnetic resonance data show clear evidence for significant helical content in water/ethylene glycol and in water/methanol mixtures at low temperatures, as well as melting behavior indicative of cooperative folding. We believe that this represents a significant step toward meeting the Paracelsus Challenge.
    Additional Material: 5 Ill.
    Type of Medium: Electronic Resource
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  • 10
    Electronic Resource
    Electronic Resource
    The effect of radical trapping reagents upon formation of poly(α-tetrahydrothiophenio para-xylylene) polyelectrolytes by the wessling soluble precursor method (1992)
    Bognor Regis [u.a.] : Wiley-Blackwell
    Journal of Polymer Science Part A: Polymer Chemistry 30 (1992), S. 2223-2231 
    Details
    ISSN: 0887-624X
    Keywords: poly(arylene vinylene) ; poly(1,4-phenylene vinylene) ; poly(para-phenylene vinylene) ; para-xylylene ; 1,4-benzoquinodimethane ; radical polymerization ; conducting polymers ; Chemistry ; Polymer and Materials Science
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology
    Notes: Molecular weights were studied by gel permeation chromatography of derivatized poly(α-tetrahydrothiophenio para-xylylene) chloride produced by aqueous or methanolic base-induced polymerization of 1,4-bis(tetrahydrothiopheniomethyl) benzene dichloride, both with and without a variety of added polymerization inhibiting agents. Efficient radical scavenging agents such as 2,2,6,6-tetramethylpiperidinoxyl and hydrogen atom donor 2,4,6-tri-tert-butylaniline reduced the degree of polymerization of the reactive α-(tetrahydrothiophenium chloride)-para-xylylene intermediate produced in this chemistry, and in some cases completely suppressed formation of high polymer. These two traps did not affect the equilibrium production of the para-xylylene by UV-Vis spectral analysis; hence they must affect the subsequent polymerization chain propagation steps in the mechanism. Electron spin resonance studies of polymerization in the presence of 0.00025 equiv of TEMPO showed disappearance of the spin label, a result consistent with a radical scavenging process. The results suggest that production of high molecular weight poly(α-tetrahydrothiophenio para-xylylene) chloride proceeds through a radical chain propagation sequence. © 1992 John Wiley & Sons, Inc.
    Additional Material: 3 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/pola.1992.080301018
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