bluuebunny/crossref_metadata_2025 huggingface dataset:
Side chain to side chain cyclization of an enkephalin analog results in loss of opioid receptor selectivity
Authors: SCHILLER PETER W. , MAZIAK LOUISE A. , LEMIEUX CAROLE , NGUYEN THI M.‐D.
Published: August 2010

The cyclic enkephalin analog H‐Tyr‐D‐L‐ys‐Gly‐Phe‐Glu‐NH2 (I) and the structurally related open chain analogs H‐Tyr‐D‐Nle‐Gly‐Phe‐Gln‐NH2 (II) and H‐Tyr‐D‐Lys(For)‐Gly‐Phe‐Abu‐NH2 (III) were tested in μ and δ opioid receptor‐representative binding assays and bioassays. Whereas both linear analogs showed a pronounced preference for μ receptors over δ receptors, the conformationally restricted cyclic peptide I was found to be unselective. This finding represents the first reported example of a peptide cyclization resulting in a loss of receptor selectivity. From this and earlier studies, it was concluded that the receptor selectivity of cyclized peptide analogs relative to that of their linear correlates may depend on the size and relative rigidity of their ring structures.
Alpha and beta momorcharins
Authors: YEUNG H.W. , LI W.W. , CHAN W.Y. , LAW L.K. , NG T.B.
Published: August 2010

Two abortifacient proteins designated α and β momorcharins were isolated from seeds of the bitter gourd Momordica charantia by a procedure involving acetone fractionation, CM‐Sepharose chromatography and gel filtration on Sephadex G‐100. The two proteins were approximately equipotent in inducing midterm abortion in mice. Hemagglutinating (lectin) activity was lacking. Homogeneity of the preparations was attested by a single band in polyacrylamide gel electrophoresis and immunoelectrophoresis. Although the two proteins had similar molecular weights as judged by SDS‐polyacrylamide gel electrophoresis, they were immunochemically distinct.
Conformation and structure of mildly heat‐treated ovalbumin in dilute solutions and gel formation at higher protein concentrations
Authors: EGELANDSDAL BJØRG
Published: August 2010

The conformation of heat‐denatured ovalbumin aggregates has been examined at several concentrations and pH values, using measurements of optical rotation dispersion (ORD), circular dichroism (CD) and viscosity. The protein was subjected to heating at relatively low temperatures, ranging from 48.5 to 76d̀; the particular temperature chosen depended on pH. The heat treatment was sufficient to remove the ability of the molecules to absorb heat on re‐heating. The denatured molecules were shown to be rather compact, i.e. not much larger than the native molecule, and to retain a significant amount of secondary structure; this was also the case for molecules present in small aggregates. It is suggested that this type of ovalbumin monomer builds three‐dimensional networks in denatured solutions at higher concentrations, and that gelation should be looked upon as arising from surface contacts between hydrated globules. The present results also imply that such globules have gelation properties which depend on whether pH is acidic or basic compared to the isoelectric point of the protein.
Backbone side chain interactions in peptides. VI
Authors: AUBRY A. , VLASSI M. , MARRAUD M.
Published: August 2010

The crystal structures of 4 dipeptides with the general formula tBuCO‐Pro‐X‐NHMe (X = His 1, His(t‐Me) 1, His(t‐Me) 1, and His+, PF6 1+) were solved by X‐ray diffraction experiments. The first 3 derivatives accommodate a βI turn conformation with the classical i + 3 ‐>i hydrogen bond. Moreover, 1 and 1 contain an additional intramolecular interaction between the His‐NH and His‐Nπ sites. The protonated dipeptide 1+ assumes a completely different conformation in which the CO(tBu) carbonyl exhibits a strong intramolecular hydrogen bond with His+‐Nπ H. The pKa of the His residue allows its existence in both the neutral and protonated states at physiological pH, and the crystal structures of 1 and 1+ are the first examples of histidine‐containing peptides investigated in the neutral and protonated states. They give some information on the possible His‐imidazole interaction modes as a function of the electrical state.
Use of intrinsic fluorescence to follow the denaturation of vicilin, a storage protein from Vicia faba
Authors: ARNTFIELD S.D. , ISMOND M.A.H. , MURRAY E.D.
Published: August 2010

Excitation of native vicilin protein at 274 nm resulted in a single emission peak at 304 nm attributed to tyrosine fluorescence. Changes in the quantum yield of the tyrosine fluorescence with addition of both urea and guanidine hydrochloride (GdnHCl) were only significant at intermediate denaturant concentrations, and not suitable for assessment of conformational change. Emission spectra for the tryptophan residues were obtained by excitation of the vicilin at 295 nm. The wavelength of maximum tryptophan emission shifted from 347 to 353 nm with the addition of denaturants (2.0 M GdnHCl or 3.0 M urea). At higher concentrations of denaturant, the quantum yield values for tryptophan also increased, reflecting a gradual change in the conformation of vicilin. The degree of polarization for both tyrosine and tryptophan fluorescence decreased significantly at denaturant concentrations slightly above those at which the protein showed the first signs of unfolding. This was further indication of a variety of conformational states for vicilin with increasing levels of denaturant.
Amino acid sequence of UP1, an hnRNP‐derived single‐stranded nucleic acid binding protein from calf thymus
Authors: MERRILL BARBARA M. , LOPRESTI MARY B. , STONE KATHRYN L. , WILLIAMS KENNETH R.
Published: August 2010

The UP1 single‐stranded nucleic acid binding protein from calf thymus (Herrick, G. & Alberts, B.M. (1976) J. Biol. Chem. 251, 2124–2132) has recently been shown to be a proteolytic fragment derived from the A1 heterogeneous nuclear ribonucleoprotein (hnRNP)(Pandolfo et al. (1985) Nucleic Acids Res. 13, 6577–6590). The NH2‐terminus of the 22 162 dalton UP1 protein appears to be blocked, which suggests that UP1 represents the NH2‐terminal two thirds of this 32 000 dalton hnRNP protein. The complete amino acid sequence for UP1 was derived from automated sequencing of peptides that were purified by HPLC from digests with trypsin, chymotrypsin, Staphylococcus aureus protease, endoproteinase Lys‐C, and cyanogen bromide. Trichloroacetic acid precipitation followed by enzymatic digestion in 2 M urea proved to be the best approach for generating UP1 peptides. By carboxymethylating after, rather than before, digestion it was possible to avoid problems associated with the insolubility of the carboxymethylated UP1. All of the resulting peptides in amounts varying from 2 to 15 nmol were coupled to aminopolystyrene prior to solid‐phase sequencing. Using these methods, no difficulties were encountered in assigning glutamic acid residues or in completely sequencing peptides that contained up to 25–30 residues. The relative ease with which the UP1 protein was sequenced, requiring only about a year to complete, and the comparatively modest amount of protein required, less than 5 mg, attests to the usefulness of water soluble carbodiimide coupling and solid‐phase sequencing for determining the primary structures of proteins. In addition to serving as a basis for determining structural relationships among various mammalian single‐stranded nucleic acid binding proteins, the amino acid sequence of UP1 reveals that the A1 hnRNP protein contains a region of internal sequence homology that apparently corresponds to two independent nucleic acid binding sites.
Proton n.m.r. spectroscopic evidence for sulfur‐aromatic interactions in peptides
Authors: LEBL MICHAL , SUGG ELIZABETH E. , HRUBY VICTOR J.
Published: August 2010

The downfield shift of the tyrosyl proton resonances and an increased chemical shift difference between the resonances for the 2′, 6′ and 3′,5′ hydrogens in a series of deamino‐oxytocin analogs modified in the disulfide bridge provide evidence for aromatic‐sulfur interactions in d6 ‐dimethylsulfoxide solutions.
Effect of pH on poppy seed 10S protein
Authors: SRINIVAS H. , RAO M.S. NARASINGA
Published: August 2010

The effect of acid pH on the 10S protein of poppy seed has been studied by ultracentrifugation, polyacrylamide gel electrophoresis, turbidity, and difference, fluorescence and circular dichroism spectra. The protein dissociated into lower molecular weight fractions in the pH range of 4.0 down to 2.5 judged by ultracentrifugation, electrophoresis and turbidity measurements. U.v. difference spectra and fluorescence measurements suggested denaturation of the protein. Thus, both dissociation and denaturation occurred down to pH 2.5. At more acid pH values (pH 1.3), reassociation and refolding probably occurred. Circular dichroism studies showed a large increase in the amplitude of the peak at pH values of 2.8 and 1.3 indicating that unordered structures are formed at acid pH values.
Synthesis of the trypsin fragment 10–25/75–88 of mouse nerve growth factor*
Authors: ROMANI S. , MORODER L. , WÜNSCH E.
Published: August 2010

Cyanogen bromide cleavage followed by trypsin digestion of mouse nerve growth factor (NGF) allowed the isolation of a double chain unsymmetrical cystine peptide of high neurotrophic activity. The presence of tryptophan residues severely limits the synthetic approaches for selective disulfide bridging. The strategy applied for such a purpose as well as the preparation of the suitably protected tetradecapeptide corresponding to sequence 75–88 as key intermediate for the synthesis of the NGF fragment 10–25/75–88 are described.
Conformations of dehydrophenylalanine containing peptides.
Authors: CHAUHAN V.S. , SHARMA ASHWANI K. , UMA K. , PAUL P.K.C. , BALARAM P.
Published: August 2010

Three tripeptides containing a central Z‐dehydrophenylalanine residue (Δz‐Phe), Boc‐L‐Phe‐Δz‐Phe‐X‐OMe (X = L‐Val 1, L‐Leu 2 and X = L‐Ala 3) have been synthesized and their solution conformations investigated by 270 MHz 1H NMR spectroscopy. In all three peptides, conformations involving the X residue NH in an intramolecular hydrogen bond were favoured in CDCl3 solutions. Studies of the nuclear Overhauser effect (NOE) provided support for a Type II β turn conformation in these peptides with Phe and Δz‐Phe occupying the i + 1 and i + 2 positions, respectively. Significantly different conformations lacking any intramolecular hydrogen bonds were observed for peptide 1 in (CD3)2SO. NOE results were consistent with a significant population of molecules having semi‐extended conformations (ø > 100°) at the Δz‐Phe residue.
Synthesis of (pGlu‐5, MePhe‐8, Sar‐9) Substance P (5–11) (DiMe‐C7) using a polyacrylamide resin and biological activity on guinea pig ileum and tracheal smooth muscle
Authors: CALAS B. , MICHELOT R. , LECAER J.P. , CAVE A. , PARELLO J. , POTIER P.
Published: August 2010

DiMe‐C7 (pGlu‐Gln‐Phe‐MePhe‐MeGly‐Leu‐MetNH2), a metabolically stable analogue of Substance P, was prepared by solid‐phase peptide synthesis using a polyacrylamide resin and a labile anchorage derived from glycolic acid. Myotropic activities in guinea pig ileum (ED50 = 4.0 ± 1.5 10‐8 M) and guinea pig trachea (ED50 = 8.6 ± 3.5 10‐8 M) are discussed in comparison with the corresponding activities of Substance P.
Isolation and sequence determination of trichorzianines A antifungal peptides from Trichoderma harzianum
Authors: HAJJI MOHAMED EL , REBUFFAT SYLVIE , LECOMMANDEUR DIDIER , BODO BERNARD
Published: August 2010

Trichorzianines A, membrane active peptides of the peptaibol class, were isolated from cultures of the mould Trichoderma harzianum. Trichorzianines A were separated into pure components by HPLC on octadecyl bonded and SiO2 phases successively. Nine trichorzianines A (IIa, IIIa, IIIb, IIIc, IVb, Vb, VIa, VIb and VII) were isolated from the complex microheterogeneous mixture. Their N‐terminal amino acid is acetylated, the C‐terminal amino alcohol is either tryptophanol or phenylalaninol, 7 to 8 of the 19 residues are α‐aminoisobutyric acid. Gas chromatography on a chiral phase showed isovaline to have the d‐configuration and all the other optically active amino acids and amino alcohols to have the l‐configuration. The amino acid sequences were determined from their positive ion FAB mass spectra which exhibited the preferential cleavage of the Aib 12‐Pro 13 amide bond as a main fragmentation. The resulting fragments subsequently underwent amide bond ruptures that generated two series of abundant acylium ions which enabled direct determination of the 1–19 sequence. The relative position of the isomeric amino acids in the sequence of trichorzianine AVII was assigned from analysis of the N‐ and C‐terminal oligopeptides yielded by its selective acidic hydrolysis. The microheterogeneity of trichorzianines A results mainly from single or multiple substitution of amino acids at the specific positions 5, 14, 16 and 19.
13C n.m.r. study of conformational equilibria in solutions of methyl N‐benzoyl‐l‐leucyl‐l‐histidinate monohydrate
Authors: HARRISON PHILIP G. , SHARPE NELSON W.
Published: August 2010

13C n.m.r. spectra for the title compound are reported in CDCl3 and also in pure and acidified DMSO solutions. In all cases, cis/trans isomerization about the amide bond occurs. For the cis isomers, severe steric compression shifts the ortho phenyl and leucyl β‐CH2 resonances to higher field. In CDCl3 solution, two imidazole conformations are present. In one, the imidazole group is freely rotating about the β‐CH2‐C(ring) bond, whilst the nitrogen‐bound proton participates in a dynamic tautomeric equilibrium, rapid on the n.m.r. time‐scale, between the two ring nitrogen atoms. In the other, the imidazole ring is protonated at N1 and held in a relatively rigid conformation by hydrogen‐bonding between N3, the water molecule, and the amide carbonyl group oxygen atom. The major imidazole species present in DMSO solution is the N1‐protonated tautomer. Small amounts of the N3 tautomer are also present, but in this solvent proton exchange between the two sites is slow on the n.m.r. time‐scale. In acidified DMSO, both nitrogen atoms are protonated.
Synthesis and conformational study of a cyclic hexapeptide analogue of somatostatin: cyclo(Phe‐D‐Trp‐Lys‐Thr‐o‐AMPA)
Authors: ELST P. VANDER , van den BERG E. , PEPERMANS H. , AUWERA L. VANDER , ZEEUWS R. , TOURWE D. , van BINST G.
Published: August 2010

The active sequence Phe7‐D‐Trp8‐Lys9‐Thr10 of somatostatin has been cyclized through o‐(aminomethyl)phenylacetic acid, a spacer molecule, designed to mimic a Gly‐Gly dipeptide containing a cis‐constrained peptide bond. The resulting analogue shows no GH‐inhibition. A 2D n.m.r. study reveals conformations different from the proposed bio‐active one and still sensitive to the medium (solvent).
Stereochemical studies on cyclic peptides. XIII. Energy minimization studies on cyclic hexapeptides having hydrogen bonds*
Authors: PAUL P.K.C. , RAMAKRISHNAN C.
Published: August 2010

The basic cyclic hexapeptide conformations which accommodate hydrogen bonded β and γ turns in the backbone have been worked out using stereochemical criteria and energy minimization procedures. It was found that cyclic hexapeptides can be made up of all possible combinations of 4 ± 1 hydrogen bonded types I, I', II and II' β turns, giving rise to symmetric conformations having twofold and inversion symmetries as well as nonsymmetric structures. Conformations having exclusive features of 3 ± 1 hydrogen bonded γ turns were found to be possible in threefold and S6 symmetric cyclic hexapeptides. The results show that the cyclic hexapeptides formed by the linking of two β turn tripeptide fragments differ mainly in (a) the hydrogen bonding scheme present in the β turn tripeptides and (b) the conformation at the α‐carbon atoms where the two tripeptide fragments link. The different hydrogen bonding schemes found in the component β turns are: 1) a β turn with only a 4 ± 1 hydrogen bond, 2) a type I or I' β turn with 4 ± 1 and 3 ± 1 hydrogen bonds occurring in a bifurcated form and 3) a type II or II' β turn having both the 4 ± 1 and the 3 ± 1 hydrogen bonds with the same acceptor oxygen atom. The conformation at the linking α‐carbon atoms was found to lie either in the extended region or in the 3 ± 1 hydrogen bonded γ turn or inverse γ turn regions. Further, the threefold and the S6 symmetric conformations have three γ turns interleaved by three extended regions or three inverse γ turns, respectively. The feasibility of accommodating alanyl residues of both isomeric forms in the CHP minima has been explored. Finally, the available experimental data are reviewed in the light of the present results.
Elephant prolactin: isolation and characterization*
Authors: LI CHOH HAO , CHUNG DAVID , BEWLEY THOMAS A. , CABRERA CLAUDIA M.
Published: August 2010

Prolactin was isolated from anterior lobes of elephant pituitary glands. It consisted of 199 amino acids with three disulfide bridges and two tryptophan residues as found in prolactin from other species. The sequence of the NH2‐terminal 28 amino acids was determined and shown homologous with the ovine hormone. In comparison with ovine prolactin, a marked difference was seen in the methionine content; the elephant hormone possessed only 18–34% lactogenic potency. The conformation of elephant prolactin was examined by zero order, second order and circular dichroism spectroscopy. The alpha helical content was estimated to be about 60%. In comparison with prolactins from other species, the second order spectra of elephant prolactin suggest that the local microenvironment for one or both tryptophan residues is somewhat different.
Synthesis of porcine neuropeptide Y(NPY) in solution
Authors: KIYAMA S. , OOI Y. , KITAGAWA K. , NAKAMURA T. , AKITA T. , KOGIRE M. , HOSOTANI R. , INOUE K. , TOBE T. , YAJIMA H.
Published: August 2010

The porcine neuropeptide Y (NPY), a 36‐residue peptide amide, was synthesized by assembling six peptide fragments followed by thioanisole‐mediated deprotection with trifluoromethanesulfonic acid in trifluoroacetic acid. β‐Cycloheptyl aspartate, Asp(OChp), was employed to suppress base‐catalyzed succinimide formation. When administered to dogs, the purified peptide (10μg/kg) caused prolonged increase of systemic arterial blood pressure and decreased pancreatic blood flow.
Conformation of sturgeon pituitary somatotropin
Authors: BEWLEY THOMAS A. , PAPKOFF HAROLD
Published: August 2010

Pituitary somatotropin (growth hormone) from the sturgeon (Acipenser guldenstadti) has been studied by zero‐order and second‐order absorption spectroscopy, as well as by circular dichroism. Difference absorption spectra have also been generated during proteolytic digestion of the hormone. The molar extinction coefficient of the native protein was found to be 15 000 ± 110 M‐1 cm‐1 at 278.5 nm. Comparison of the conformations of sturgeon somatotropin and somatotropins isolated from several mammalian species, including bovine and human, indicates a close relationship between these molecules. Such similarities may be related to the relatively high biopotency of this fish hormone in mammalian assay systems.
Fourth polymorph of [Phe 4 Val 6] antamanide (pentahydrate)
Authors: KARLE ISABELLA L. , WIELAND THEODOR
Published: August 2010

The cyclic decapeptide antamanide and the synthetic, biologically active analog [Phe4 Val6] antamanide (cyclic[ValProProPhePhe]2) crystallize in various crystal forms as a function of the solvent. The present crystalline polymorph obtained from acetone/water (also from ethanol/water and DMSO/water) crystallizes in space group P212121 with a = 20.194 (30) Å, b = 21.118 (31) Å, c = 16.132 (25) Å and four molecules of peptide in the unit cell. There are five cocrystallized water molecules per peptide molecule, of which four water molecules are intrinsic to the peptide molecule. Although the molecular packing is entirely different in each of the polymorphs, the conformation of the peptide molecule, including the intrinsic water molecules, is very similar in all the polymorphs.
Ontogeny of the bovine neurohypophysial hormone precursors.
Authors: LEVY B. , ROUILLE Y. , CHAUVTE M.T. , CHAUVET J. , ACHER R.
Published: August 2010

Neurohypophysial hormone precursors are small proteins processed into several fragments during axonal transport from hypothalamus to neurohypophysis. From 3‐month‐old fetal bovine pituitaries the three fragments of vasopressin precursor, arginine vasopressin, MSEL‐neurophysin and copeptin, and the two fragments of oxytocin precursor, oxytocin and VLDV‐neurophysin, have been isolated and characterized. These polypeptides are identical to those previously identified in the late fetus (7–9 months old) and in the adult. It is concluded that the same genes are expressed during fetal and adult lifes, the vasopressin gene appearing roughly four times more active than the oxytocin gene in the early fetus. Vasotocin, mesotocin and additional neurophysin have not been detected in the early fetus.
Synthesis of a cyclic retro analogue of somatostatin suitable for photoaffinity labelling
Authors: KESSLER H. , HAUPT A. , FRIMMER M. , ZIEGLER K.
Published: August 2010

Cyclic somatostatin analogues containing the modified retro sequence of the amino acids Phe7 to Phe11 of the natural compound have been found to exhibit high activity for cytoprotection of rat hepatocytes against cell poisons such as phallotoxins and galactosamine. Cyclo(‐Phe(p‐NH(1‐14C)Ac)‐Thr‐Lys(CO(p‐N3)C6H4)‐Trp‐Phe‐d‐Pro‐), a photoreactive and radioactive analogue of one of the most active cyclohexapeptides, was synthesized by a combination of solid phase technique and classical solution peptide synthesis. This peptide labels the same proteins in rat liver cell membrane that are modified by photolysable derivatives of bile acids, phalloidin and antamanide.
Synthesis of N‐(4‐aminobenzoyl)‐γ‐oligo (L‐glutamic acid)s*
Authors: KRZYZANOWSKI LESZEK , RZESZOTARSKA BARBARA
Published: August 2010

N‐(4‐aminobenzoyl)‐γ‐oligo (l‐glutamic acid)s (6) containing from two to six glutamic residues have been prepared in solution using Nα‐Boc‐α‐Bzl protections and isobutyl‐chlorocarbonate activation. Key steps in the synthesis were the coupling of γ‐oligo(α‐benzyl l‐glutamate) benzyl esters (1) with N‐(4‐benzyl‐oxycarbonylaminobenzoyl)‐l‐glutamic acid α‐benzyl ester (4) to blocked precursors of N‐(4‐aminobenzoyl)‐γ‐oligo (l‐glutamic acid)s (5) and catalytic hydrogenolysis of 5 to 6. Elaboration of the required oligo γ‐l‐glutamate chains (1) was achieved step by step beginning with the coupling of glutamic acid dibenzylester with N‐(t‐butoxycarbonyl)‐l‐glutamic acid α‐benzyl ester (2) to 3 followed by selective removal of the Boc from 3 with HCl‐dioxane followed by coupling with 2.
Formation of a cyclic imide in aspartyl or asparaginyl glycyl peptides induced by heating in the dry state*
Authors: LUO SIQUAN , LIAO CHU‐XIONG , McCLELLAND JOHN F. , GRAVES DONALD J.
Published: August 2010

A common product was identified by HPLC from the heating of, α and β isomers of, aspartyl glycine in the dry state. Fast atom‐bombardment (FAB) mass spectrometry and Fourier transform infrared photoacoustic spectroscopy (FTIR‐PAS) showed that the product is a cyclic imide. Cyclic imide formation also was shown to occur with the tripeptides Gly‐Asp‐Gly and Gly‐Asn‐Gly. The cyclization of aspartyl peptides depends upon protonation of the β‐carboxyl group and can be promoted by acidification or by heating the NH4+ salt in the dry state.
Purification and primary structure of ostrich pancreatic polypeptide
Authors: LITTHAUER DEREK , OELOFSEN WILLEM
Published: August 2010

Pancreatic polypeptide has been isolated from ostrich pancreas by gel filtration, ion exchange chromatography and high pressure liquid chromatography. The ostrich peptide contains 36 amino acids and has an amino acid composition similar to pancreatic polypeptide of other avian species. The primary structure of ostrich pancreatic polypeptide differs from that of the chicken peptide only at residues 3 and 18 where the ostrich peptide contains an alanine and a valine residue compared to the serine and isoleucine residues found in those positions in the chicken peptide.
Metal‐ligated amino acids and peptides as substrates for proteases
Authors: BAGGER SVEN
Published: August 2010

Amino acids and peptides carrying a pentaamminecobalt(III) group at the carboxyl terminal have been prepared. It is shown that trypsin and papain accept such compounds as substrates provided the metal complex group is not too close to the enzyme‐susceptible peptide bond. The possible applicability of this novel type of substrates in enzymatic peptide synthesis is discussed.
Reduction and reoxidation of the neurotoxin II from the scorpion Androctonus australis Hector
Authors: SABATIER J.‐M. , DARBON H. , FOURQUET P. , ROCHAT H. , VAN RIETSCHOTEN J.
Published: August 2010

Reoxidation of the totally reduced scorpion neurotoxin II from Androctonus australis Hector (four disulfide bridges) has been investigated. The totally reduced toxin was highly insoluble in neutral and alkaline conditions, which prevented the use of the usual air oxidation process for renaturation. We tested a new method in which the reduced molecules were first solubilized in 10% (v/v) acetic acid and then oxidized by air through dialysis against a series of buffers with a slow pH gradient from 2.2 to 7.0 or 8.0. In this system, up to 95% of the protein was recovered in solution. Addition of reduced and oxidized glutathione accelerated refolding and also permitted a better recovery of fully active peptide as measured by both toxicity to mice and ability to displace 125I radiolabeled toxin II from its binding site on rat brain synaptosomal fractions. The reoxidation reaction could also be monitored directly by high pressure liquid chromatography. A strong effect of guanidine hydrochloride concentration as well as of temperature was observed both on the solubility of the reoxidation intermediates and on the refolding pathway. Finally, the method used, i.e. dialysis reoxidation with a pH gradient from 2.2 to 8.0 in 0.1 M sodium phosphate, 0.1 M sodium chloride, 20 mM guanidine hydrochloride, 1 mM oxidized and reduced glutathione allowed regeneration in high yield (70%) of a reoxidized toxin form indistinguishable from the native toxin. A minor stable and inactive molecular species (about 30%) showing a difference in mobility by electrophoresis was also detected.
Binding of C‐terminal segments of neuropeptide Y to chicken brain
Authors: PERLMAN MICHAEL O. , PERLMAN JANET M. , ADAMO MARTIN L. , HAZELWOOD ROBERT L. , DYCKES DOUGLAS F.
Published: August 2010

The C‐terminal pentapeptide amide segment of neuropeptide Y (NPY) binds specifically to chicken brain membrane preparations. The contribution of each residue of the C‐terminus to this binding has been investigated through the synthesis and evaluation of a series of pentapeptide analogs. The binding of these molecules is strongly dependent on the presence of certain functional groups, in particular the guanidinium group of Arg‐35 and the C‐terminal aromatic amide function. The significance of these results for the binding to chicken brain tissue of NPY, pancreatic polypeptide and other potentially related neuropeptides is discussed.
An acid‐labile anchoring linkage for solid‐phase synthesis of C‐terminal peptide amides under mild conditions*
Authors: ALBERICIO FERNANDO , BARANY GEORGE
Published: August 2010

A series of polymer‐supported benzylamides substituted with one to three alkoxy groups in the ring positions were prepared and shown to give carboxamides upon treatment with acid. Based on the initial screening, the bis(o‐methoxy)‐p‐alkoxybenzylamide anchoring linkage was selected for a detailed evaluation of its suitability for solid‐phase synthesis of C‐terminal peptide amides. The handle derivative 5‐[(2′ or 4′)‐Fmoc‐aminomethyl‐3′,5′‐dimethoxyphenoxy]valeric acid (1) was prepared in seven facile steps [purification of intermediates unnecessary; overall yield 15% for crystalline product, which is a mixture of positional isomers], and was quantitatively coupled onto amino group‐containing supports by use of N,N'‐dicyclohexylcarbodiimide plus 1‐hydroxybenzotriazole in N,N‐dimethylformamide. Stepwise elaboration of peptide chains proceeded smoothly with both Nα‐9‐fluorenyl‐methyloxycarbonyl (Fmoc) and Nα‐dithiasuccinoyl (Dts) amino acids, and final cleavage of tert.‐butyl side‐chain protecting groups and of the anchoring linkage occurred readily in trifluoroacetic acid–dichloromethane (7:3) at 25°. The methodology was demonstrated by the syntheses of H‐Trp‐Asp‐Met‐Phe‐NH2 (tetragastrin) and H‐Tyr‐Gly‐Gly‐Phe‐Met‐NH2 (methionine‐enkephalinamide), both with high yields and purities.
Structural identity of the subunits of pigeon liver malic enzyme
Authors: KAM PUI‐LING , LIN CHING‐CHUN , CHANG GU‐GANG
Published: August 2010

Pigeon liver malic enzyme was found to have arginine, alanine, and tyrosine as the only N‐terminal, N‐1, and N‐2 amino acids, respectively. Hydrolysis of the reduced and carboxymethylated malic enzyme by carboxypeptidase A yielded quantitative evidence for the following C‐terminal sequence: ‐Leu‐(Phe‐Ala)‐Ile‐Leu‐COOH. Fifty‐five trypsin‐digested peptides were separated by HPLC, in accordance with the arginine and lysine contents of each subunit. This more direct structural evidence strongly supports the conclusion that pigeon liver malic enzyme is composed of four chemically identical subunits.
Comparative conformation‐activity relationships for hormonally‐ and centrally‐acting TRH analogues
Authors: WARD D.J. , FINN P.W. , GRIFFITHS B.C. , ROBSON B.
Published: August 2010

Empirical energy calculations have been applied to a series of thyrotrophin‐releasing hormone (TRH) analogues in an attempt to incorporate the conformational parameter into traditional Quantitative Structure‐Activity Relationships (QSAR). A search was made of the potential surface of each analogue using a SIMPLEX technique coupled with a model representing solvent effects as a dielectric continuum. Conformers of TRH can be described in terms of the distance between the pyroglutamyl, imidazole and prolyl rings, and presented as two‐dimensional descriptor coordinates using multidimensional scaling. This process was repeated for stable and metastable conformers of all the TRH analogues investigated and correlated with experimental data on receptor affinity, relative stability and biological potency of the analogues. The results obtained suggest that modifications to the TRH structure can lead to greater potency in addition to improved stability, and that providing the appropriate theoretical approach is taken, it is possible to apply QSAR‐based design procedures to conformationally‐flexible drugs.
Molecular mechanics investigation of the flexibility of some cyclic peptides
Authors: ROMANOWSKA KRYSTYNA , KOPPLE KENNETH D.
Published: August 2010

Conformation space near the crystal conformations of proline‐containing cyclic octapeptides and cyclic hexapeptides of C2 sequence symmetry, e.g. cyclo‐(Gly‐Pro‐d‐Phe)2 and cyclo‐(d‐Ala‐Gly‐Pro‐d‐Phe)2, was explored using molecular mechanics. Conformations found in crystals were energy minimized, distortions were introduced by systematically fixing backbone dihedral angles at individual residues, and nearby energy‐minimized conformations were then located. Interatomic distances and dihedral angles were examined in the conformations within a few kilocalories of the most stable conformation. A common form of flexibility was found to involve libration of amide planes. Among the peptides examined, the cyclic hexapeptides were found to have greater freedom than the cyclic octapeptides, and cyclo‐(d‐Ala‐Gly‐Pro‐d‐Phe)2 was found to be more rigid than cyclo‐(d‐Ala‐Gly‐Pro‐Phe)2.
Synthesis of cyclic tryptathionine peptides
Authors: ZANOTTI GIANCARLO , BEIJER BARBRO , WIELAND THEODOR
Published: August 2010

The helicity of the tryptathionine moiety of the phallotoxins has been recognized by comparison with cyclic tryptathionine tripeptides. In order to investigate the influence of the configuration of the component amino acids on the conformation of the cyclic peptides, six analogue thioether tripeptides containing L‐ and D‐alanine and L‐ and D‐cysteine, respectively, have been synthesized. The CD spectra of the peptides are very similar to each other, showing mirror images of the CD of phalloidin and, therefore, negative helicity. The spectra of the D‐cysteine containing compounds differ from the L‐cysteine containing compounds by their weakly positive ellipticity values around 270 nm. The cyclization reaction of Boc‐Hpi‐D‐Ala‐D‐Cys(STrt)OCH3, along with the cyclic tripeptide, afforded a cyclic hexapeptide by dimerization. The CD spectrum of the dimer is very similar to that of phalloidin, thus pointing to a positive helicity of its two tryptathionine moieties. The dimeric thioether peptide forms a rather strong complex with Cu2+ ions.
Ethoxyformylation of histidine residues in equine growth hormone
Authors: FUKUSHIMA J.G. , CASCONE O. , SANTOMÉ J.A. , BONINO M.J. BISCOGLIO DE JIMENEZ
Published: August 2010

Reactivity of histidine residues in equine growth hormone to ethoxyformic anhydride was studied. The existence of two kinetically different sets was demonstrated: one of them including only the slow reacting histidine 169 (k = 0.164min‐1) and the other containing fast reacting histidines 19 and 21 (k = 0.892 min‐1). A correlation between the decrease in the capacity to compete with125I‐labeled hormone for rat liver binding sites and the degree of ethoxyformylation of the fast group was found. Circular dichroism studies indicated no significant conformational changes in the protein with all three residues modified. These results fully agree with those obtained for bovine growth hormone which is further evidence supporting the vinculation of histidines 19 and/or 21 with the binding site of these hormones to their specific receptors.
Conformational analysis of a cyclic thymopoietin‐analogue by 1H n.m.r. spectroscopy and restrained molecular dynamics simulations
Authors: LAUTZ J. , KESSLER H. , BOELENS R. , KAPTEIN R. , VAN GUNSTEREN W.F.
Published: August 2010

The internuclear distances of the cyclic thymopoietin derivative c[D‐Val‐Tyr‐Arg‐Lys‐Glu] have been determined using two‐dimensional nuclear Overhauser n.m.r. spectroscopy. These distances are used as constraints for a restrained Molecular Dynamics (MD) simulation. The two starting structures used for the calculations consist of a β and γ turn for model 1 and two γ turns for model 2. The rms difference in atomic positions of the two conformations is 0.242 nm. They converge during the restrained MD simulation to the same final structure. The positional rms difference of the time averaged (5–14ps) conformations is 0.011 nm. The hydrogen bond pattern is similar to that of model 1, but in addition we find three more γ turns. The vicinal NH‐Cα H couplings agree well with those calculated from the time averaged structures.
Applications of ammonium formate‐catalytic transfer hydrogenation.
Authors: ANWER MOHMED K. , PORTER RICHARD A. , SPATOLA ARNO F.
Published: August 2010

Dehalogenation of chlorine bound to aromatic nuclei has been achieved using ammonium formate‐catalytic transfer hydrogenation (AF‐CTH) techniques. The use of deuterated ammonium formate as the transfer agent in the CTH process when performed in a deuterated solvent medium results in the predominant formation of the labeled product. Thus, ND4+ DCOO‐ ‐ CTH of [D‐Ala2, p‐ClPhe4]‐Leu‐enkephalin in 80% CD3COOD/D2O yielded [D‐Ala2, Phe (4D)4]‐Leu‐enkephalin. Levels of D‐incorporation were measured by fast atom bombardment‐mass spectrometry (FAB‐MS) and/or by 13C n.m.r. spectroscopy. Using D‐labeled and unlabeled ammonium formate in stoichiometrically similar reactions, it was concluded that the hydrogenation exhibited a primary kinetic isotope effect. Reactant and product concentrations were determined by amino acid analysis and reversed phase HPLC.
Cyclopropane amino acid ester dipeptide sweeteners
Authors: MAPELLI CLAUDIO , NEWTON M. GARY , RINGOLD C.E. , STAMMER C.H.
Published: August 2010

A series of esters of L‐aspartyl‐1‐aminocyclopropane carboxylic acid has been prepared and their sweet tastes determined. The sweetest ester prepared was about 300 times sweeter than sucrose. An attempt to use basic conditions during preparation of the dipeptide allyl ester led to succinimide formation of the aspartyl peptide even though the β‐carboxyl group was protected by a t‐butyl ester function. The X‐ray structure of the propyl ester (1c) was determined and its conformation is discussed.
Comparison of the interaction of methionine and norleucine‐containing peptides with phospholipid bilayers
Authors: EPAND RICHARD M. , RAYMER KAREN E.
Published: August 2010

Norleucine is a structural analog of methionine with a methylene group replacing the thio ether. Despite the close structural similarity of these two amino acids, norleucine‐containing peptides have markedly different behaviour with phospholipids compared with methionine‐containing peptides. For example, HCO‐L‐Ahx‐L‐Leu‐L‐Phe‐OMe behaves as a hydrophobic peptide when mixed with dimyristoylphosphatidylcholine. This peptide lowers the enthalpy of the lipid phase transition. The effect is independent of the rate of heating. With the homologous peptide, HCO‐L‐Met‐L‐Leu‐L‐Phe‐OMe, the results are markedly dependent on scan rate with a higher enthalpy observed at faster scan rates. Only at a scan rate of 0.2 K min‐1 do the two peptides approach similar behaviour. The higher enthalpy observed for samples with the methionine peptide at higher scan rates can be explained assuming that the peptide aggregates at low temperature. As the phase transition temperature is approached, the more hydrophilic methionine peptide partitions more slowly into the membrane than the norleucine peptide. Partitioning of the peptides between aqueous and lipid phases was measured at 37° by centrifuging down the lipid‐bound fraction. At a peptide concentration of 15 μM and a lipid concentration of 1.4 mM, 89% of the HCO‐L‐Ahx‐L‐Leu‐L‐PheOMe and 97% of the HCO‐L‐Met‐L‐Leu‐L‐PheOMe remained in the supernate, indicating a greater tendency of the norleucine‐containing peptide to partition into the lipid phase. The peptides Ac‐L‐Phe‐L‐Met‐L‐Arg‐L‐Phe‐NH2 and Ac‐L‐Phe‐L‐Ahx‐L‐Arg‐L‐Phe‐NH2 are readily soluble in water. These peptides were deposited with the lipid from a solution in chloroform/methanol. After repeated scanning and/or incubation at the phase transition temperature the effect of the peptide in raising the phase transition temperature of the lipid was lost. Thus these peptides were no longer incorporated into the bilayer after these treatments. The loss of the alterations of the phase transitions caused by these peptides was observed upon immediate rescanning of the methionine‐containing samples while for the norleucine‐containing ones, incubation for several hours at the phase transition was required. Our results demonstrate that norleucine‐containing peptides partition more rapidly into the lipid phase while the methionine peptides are more rapidly extracted from the lipid into the aqueous phase. The results emphasize the importance of determining the effect of scan rate as a criterion of the attainment of equilibrium in lipid‐peptide mixtures. We demonstrate how relatively minor changes in structure can alter the behaviour of peptides with lipids. Our results suggest that the pharmacodynamics of peptides in which norleucine has replaced methionine will be altered.
Influence of secondary structure of polyglycine on some conformational properties
Authors: DOLCET C. , TARAZOAN M.P.
Published: August 2010

Values of four conformational properties, namely unperturbed dimension < r2>0, dipole moment < μ2 >, mean squared optical anisotropy < γ2 >, and molar Kerr constant < mK >, have been calculated for polyglycine chains allowing several combinations of the secondary structure with the aim of studying the dependence of these magnitudes on the secondary structure of the chain. Two different approaches to the secondary structure have been used. In the first, chains with all their units in a given conformation (random coil, α‐helix or β‐sheet) are interrupted at several positions by one unit in a different conformation. In the second, chains with varying composition of two conformations α‐helix/β‐sheet and β‐sheet/random coil were allowed and the results obtained compared with previous work for α‐helix/random coil chains.
Crystal structure and conformation of polypeptides: L‐leucylglycylglycylglycine
Authors: SRIKRISHNAN T. , PARTHASARATHY R.
Published: August 2010

Crystals of L‐leucylglycylglycylglycine, LGGG (C12H22N4O5), grown from an ethanol‐water solution, are orthorhombic, space groups P212121, with unit cell dimensions (at 22 ± 3°) a = 9.337(1), b = 10.995(1), c = 15.235(1)Å, v = 1563.4 Å3, Z = 4 with a density of Dobs= 1.29 g‐cm‐3 and Dcalc= 1.279 g°cm‐3. The crystal structure was solved by the application of direct methods and refined to an R value of 0.029 for 1018 reflections with I ± 2s̀. The molecule exists as a zwitterion in the crystal. The trans peptide backbone takes up a folded conformation at the middle glycylglycyl link accompanied by a significant nonplanarity up to Δω of 8° at the middle peptide and is relatively more extended at the two ends. The molecules are linked together intermolecularly in an infinite sequence of head to tail 1–4′ hydrogen bonds, as is typical of charged peptides. It is interesting to note that while glycylglycylglycine takes up an extended β‐sheet conformation, addition of Leu to the N‐terminal results in a bent conformation.
Stereochemistry of peptides containing a thioacyl group
Authors: LA COUR TROELS F.M.
Published: August 2010

Endothiopeptides are oligopeptides in which one or more oxoacyl moieties in peptide groups are replaced with thioacyl moieties. The stereochemical consequences of this replacement are discussed and compared with results from X‐ray crystallographic determinations of small oligopeptides containing one or two thiopeptide groups. The analysis shows that the conformations normally observed around Cα carbon atoms in proteins are also stereochemically favourable for peptides in which a thiopeptide group has been introduced. Limitations are only present in cases where the thiopeptide is part of structural elements like helices and β‐sheets. This opens the possibility of using one or more thiopeptide groups as functional modifiers of biologically active oligopeptide and protein molecules.
Conformational preferences and self‐association modes of two diastereomeric statine derivatives
Authors: TONIOLO CLAUDIO , VALLE GIOVANNI , BONORA GIAN MARIA , LELJ FRANCESCO , BARONE VINCENZO , FRATERNALI FRANCA , CALLET GEORGES , WAGNON JEAN , NISATO DINO
Published: August 2010

The conformational preferences and self‐association modes of the two diastereomeric N‐acetyl, methylamides of 3‐hydroxy, 4‐amino, 6‐methylheptanoic acid (statine) with (R, S) and (S, S) configurations at the 3‐hydroxy and 4‐amino carbons, respectively, have been determined in solution as well as in the solid state by infrared absorption, 1H nuclear magnetic resonance, and X‐ray diffraction. Conformational energy computations have also been performed in parallel. In the crystal state, the change in chirality of the hydroxyl group induces different intermolecular H‐bonding schemes in the (R, S) isomer compared to the two structurally distinct molecules in the asymmetric unit of the (S, S) isomer. Different propensities to self‐aggregate are seen in solvents of low polarity. In solvents of high polarity, however, the molecules of both isomers are largely solvated, while still keeping some local conformational restriction. Conformational energy computations indicate that in vacuo the two diastereomers exhibit different flexibility, and a preferred conformation with a different type of intramolecular H‐bond.
Conformation similarities of ricin A‐chain and trichosanthin
Authors: KUBOTA SHIGEO , YEUNG H.W. , YANG JEN TSI
Published: August 2010

The conformation of ricin A‐chain from castor bean was studied by circular dichroism at pH 4.7, 7 and 9 and compared with that of trichosanthin from the Chinese herb Tianhuafen. The CD spectra of ricin A‐chain and trichosanthin were nearly identical at each of the three pHs. Analysis of the data indicated that, like trichosanthin, ricin A‐chain had about 29%α‐helix and 42%β‐sheet but no β‐turn. However, there was a subtle difference in the CD spectra in 20 mm sodium dodecyl sulfate, the addition of which at pH 7 slightly increased the helicity and decreased the content of β‐sheet of ricin A‐chain in contrast to a larger increase in helicity at the expense of β‐sheet for trichosanthin, thus indicating a different stability against the surfactant. Native ricin A‐chain and trichosanthin had about the same amount of secondary structure, which supports the belief that a high degree of sequence homology of the two proteins [Zhang & Wang (1986) Nature 321, 477–478] may lead to a conformational similarity between them, even though the two proteins are not taxonomically related.
Automated synthesis and use of N‐chloroacetyl‐modified peptides for the preparation of synthetic peptide polymers and peptide‐protein immunogens
Authors: LINDNER WOLFGANG , ROBEY FRANK A.
Published: August 2010

A method to incorporate N‐chloroacetyl moieties at the amino termini of synthetic peptides using a standard program with an automated peptide synthesizer has been developed. The N‐chloroacetyl‐modified peptides react well with sulfhydryl containing proteins such as 4‐mercaptobutyrimide‐modified bovine serum albumin to form stable protein‐peptide conjugates. By incorporating cysteine into the synthetic peptide, autopolymerization or cyclization of the synthetic peptide occurs by reaction of the free sulfhydryl with the chloroacetyl group. N‐Chloroacetyl‐derivatized peptides may be useful as reagents for potential peptide immunogens and vaccines.
Propensity for spontaneous succinimide formation from aspartyl and asparaginyl residues in cellular proteins
Authors: CLARKE STEVEN
Published: August 2010

One mechanism for the spontaneous degradation of polypeptides is the intramolecular attack of the peptide bond nitrogen on the side chain carbonyl carbon atom of aspartic acid and asparagine residues. This reaction results in the formation of succinimide derivatives and has been shown to be largely responsible for the racemization, isomerization, and deamidation of these residues in several peptides under physiological conditions (Geiger, T. & Clarke, S. J. Biol. Chem. 262, 785–794 (1987)). To determine if similar reactions might occur in proteins, I examined the sequence and conformation about aspartic acid and asparagine residues in a sample of stable, well‐characterized proteins. There did not appear to be any large bias against dipeptide sequences that readily form succinimides in small peptides. However, it was found that aspartyl and asparaginyl residues generally exist in native proteins in conformations where the peptide bond nitrogen atom cannot approach the side chain carbonyl carbon to form a succinimide ring. These orientations also represent energy minimum states, and it appears that this factor may account for a low rate of spontaneous damage to proteins by succinimide‐linked reactions. The presence of aspartic acid and asparagine residues in other conformations, such as those in partially denatured, conformationally flexible regions, may lead to more rapid succinimide formation and contribute to the degradation of the molecule. The possible role of isoimide intermediates, formed by the attack of the peptide oxygen atom on the side chain carboxyl group, in protein racemization, isomerization, and deamidation is also considered.