International journal of peptide and protein research最新文献

The adipoyl- and suberoyl-linked bis(Ala) peptides have an extended backbone between the two C alpha atoms in each molecule. They self-assemble, through intermolecular hydrogen bonds and stacking of parallel strands, into highly ordered modified beta-sheet-like structures. Crystals data for adipoyl bis(Ala)diester are as follows: C14H24N2O6, monoclinic space group P2(1), a = 4.900(1), b = 29.093(10), c = 6.021(2) A, beta = 104.20(2) degrees, R = 0.053 for 1100 data > 3 sigma (F); for suberoyl bis(Ala)diester: C16H28N2O6, monoclinic space group P2(1), a = 4.887(2), b = 32.650(9), c = 6.004(2) A, beta = 103.79(3), R = 0.070 for 1065 data > 3 sigma (F).

Complete proton resonance assignments of the naturally occurring mu-selective dermorphin (H-Tyr-D-Ala-Phe-Gly-Tyr-Pro-Ser-NH2) and delta-selective deltorphin-I (H-Tyr-D-Ala-Phe-Asp-Val-Val-Gly-NH2) were carried out by two-dimensional 1H-NMR techniques to investigate the conformational features in the membrane-mimetic micelles of perdeuterated dodecylphosphocholine. Fifty possible three-dimensional structures for respective peptides were generated by means of distance geometry calculations, all of which satisfy the proton-proton distances derived from NOE measurements within the allowable range, and 25 of them were subjected to the molecular dynamics simulations for 10 ps, in which the NOE distances were included as the energetic constraints. Although conformers simulated for dermorphin showed relatively large conformational variations because of the limited NOE data, most of them were characterized as an entirely folded structure bent at the Gly4 residue, where each of the N- and C-terminal tetrapeptides took an extended conformation. On the other hand, most conformations of deltorphin-I showed the common feature that the N-terminal Tyr-D-Ala-Phe-Asp and C-terminal Val-Val-Gly-NH2 sequences took respective folded conformations, and these were almost at right angles on the border of the Asp-Val sequence. These conformational characteristics are discussed in terms of the possible relationship with the mu/delta-opioid receptor selectivity.

The backbone-modified tripeptide Bz-Aib-NHCOCO-Aib-OMe, in which the central C alpha has been replaced with CO, self-assembles in the solid state into highly ordered two-dimensional arrays through MeOH mediated intermolecular stacking of dimeric 'disk' modules formed by an antiparallel beta-sheet-type arrangement of tripeptide molecules. The C alpha(C'O)N(C'O)(C')NC alpha segment is nearly coplanar (average deviation from a mean least-square plane is +/- 0.037 A). The oxalyl moiety forms two pseudo-C5 intramolecular hydrogen bonds. Crystal parameters are as follows: C18H23N3)6.CH3OH, triclinic space group P1-, alpha = 9.796(4), b = 10.348(3), c = 11.836(4) A, alpha = 78.28(3), beta = 73.72(3), gamma = 69.45(3), R = 0.082 for 1747 reflections measured with [formula: see text].

Several phosphoserine, phosphothreonine and phosphotyrosine synthons suitable for the stepwise synthesis of phosphopeptides were prepared. Treatment of methylthiomethyl (MTM) esters of either Z-, Boc-, Allocserine and threonine with phosphochloridate in pyridine followed by MgBr2 cleavage of MTM in diethyl ether afforded the title compounds in good yield. Thiophosphoserine and phosphotyrosine synthons were also obtained by the phosphoramidite method using di-(2,2,2-trichloroethyl)-N,N-diisopropylphosphoramidite and MCPBA as oxidizing reagent. Trichloroethyl proved valuable as phosphate protecting group especially in phosphotyrosine derivatives owing to its stability in acidic conditions. These synthons were involved in the liquid-phase synthesis of several phospho and/or thiophosphopeptides related to either src-protein kinase or rat liver pyruvate kinase.

A convenient enantioselective synthesis of p-hydroxymethyl-L-phenylalanine was developed which produces a 4/1 ratio of L/D enantiomers resulting from a chiral phase-transfer-catalyzed alkylation. This amino acid was coupled into the p56(1)ck tyrosine kinase substrate Ac-Leu-Pro-Tyr-Ala-NHCH3 as a replacement for Tyr and can subsequently be elaborated into a variety of potential tyrosine kinase inhibitor designs of general structure Ac-Leu-Pro-AA-Ala-NHCH3, wherein AA is an unnatural amino acid. The contaminating D enantiomer was readily removed after coupling to L-Ala-NHCH3 of this sequence. The utility of the p-hydroxymethyl functionality in an efficient divergent synthetic strategy leading to various inhibitor designs is illustrated with the synthesis of Ac-Leu-Pro-AA-Ala-NHCH3, wherein AA is p-(R,S-hydroxyphosphonomethyl)-L-phenylalanine.

The influence of low temperature and high viscosity on the conformation of somatostatin and two of its analogues was investigated by 1H NMR in solution. The conformation of native somatostatin, a cyclic octapeptide agonist DC13-116 and a linear octapeptide agonist were compared in ethylene glycol at 303 K and in methanol at low temperature. The first goal of this study was to investigate if either low temperature or high viscosity is the more important for the reduction of the conformational freedom. Secondly we wanted to compare the amount of information concerning the conformation present in both solvents. A larger amount of NOESY cross-peaks is observed in ethylene glycol at room temperature compared to methanol at low temperature. This indicates that the raising of the viscosity is a more important factor in reducing the flexibility of peptides than the lowering of the temperature.

A conformation analysis of tetrapeptide in DMSO was carried out by the Monte Carlo simulation including the solvent effects in the energy function. The lowest energy conformations in the zwitterionic and cationic states are a similar compact form, in which the two aromatic rings are close to each other. The distance distributions between the specified atoms in the molecule show that most conformations in the local energy minima are similar to this folded conformation, although there is another type of conformation of interest, which has an extended form with the two separate aromatic rings. Such results are consistent with the NMR and CD experiments, and suggest that the compact form of tetragastrin is essential as a biological active conformation. The folded structure is stabilized by the solute-solvent interaction. In particular, the nonbonding interactions between the solute and solvent molecules cause a folding effect on the molecular conformation in total. This study is a first case taking account of the solvent effect into the conformation analysis of tetragastrin.

We recently described a two-step enzymatic semisynthesis of the superpotent analog of human growth hormone releasing factor, [desNH2Tyr1,D-Ala2,Ala15]-GRF(1-29)-NH2 (4), from the precursor, [Ala15,29]-GRF(4-29)-OH (1). C-Terminal amidation of 1 to form [Ala15]-GRF(4-29)-NH2 (2) was achieved by carboxypeptidase-Y-catalyzed exchange of Ala29-OH for Arg-NH2. The target analog 4 was then obtained by acylation of segment 2 with desNH2Tyr-D-Ala-Asp(OH)-OR (3) (R = CH3CH2- or 4-NO2C6H4CH2-) catalyzed by the V8 protease. In this paper we report on the use of the recently isolated Glu/Asp-specific endopeptidase (GSE) from Bacillus licheniformis, which is shown to be an efficient catalyst for the segment condensation of 2 and 3. GSE is more stable than the V8 protease under the conditions employed (20% DMF, pH 8.2, 37 degrees C). The extent of conversion of 2 into 4 is limited by proteolyses at Asp3-Ala4 and Asp25-Ile26. However, this proteolysis is virtually eliminated by use of the appropriate ester leaving group, R. A systematic study of the kinetics of the GSE-catalyzed segment condensations of 2 and a series of tripeptide esters, desNH2Tyr-D-Ala-Asp(OH)-OR (3) [R = CH3CH2- (3a), CH3- (3b), ClCH2CH2- (3c), C6H5CH2- (3d), 4-NO2C6H4CH2- (3e)] revealed that rate of aminolysis versus proteolysis, and hence the conversion of 2 into 4, increase with increasing specificity (Vmax/Km) of GSE for the tripeptide ester.(ABSTRACT TRUNCATED AT 250 WORDS)

Perloza beaded cellulose was functionalised by a cyanoethylation/reduction procedure to give aminopropyl Perloza. Fmoc-amino acids were anchored to aminopropyl Perloza beaded cellulose via the TFA labile 4-oxymethylphenoxyacetyl (HMPA) linker. Using Fmoc-aminoacyl-4-oxymethylphenoxyacetyl-2,4-dichloro-phenyl esters, all 20 amino acids were anchored at substitution levels ranging from 0.37 to 0.65 mmol/g. Fmoc-amino acids were also anchored using the peptide-amide linker 4-[(R,S)-1-[1-(9H-fluoren-9-yl)-methoxycarbonylamino - (2',4'-dimethoxybenzyl]phenoxyacetic acid. The Fmoc-aminoacyl resins were used for SPPS using Fmoc chemistry. SPPS was carried out using either an LKB Biolynx 4175 low-pressure pumped column continuous-flow peptide synthesiser or an ABI 430A automated vortexing batchwise instrument. Comparison of peptides made using each synthesiser showed little difference in quality of the crude peptides. Different Fmoc-amino acid activation methods (DIC/HOBt/DMF, HBTU, DIC/HOBt/DCM) were found to be equally useful with Perloza. Peptides were cleaved using TFA plus scavengers; however, the TFA-swollen resin was not readily separated from the TFA/peptide solution by simple filtration. Therefore alternative cleavage workup procedures were used with Perloza. Peptides were purified by HPLC and characterised by HPLC and amino acid analysis, and in some cases by FAB-MS. Successful syntheses ranged from 5 to 34 amino acids in length. Some of the peptides were also synthesized using a polystyrene support and standardised (ABI Fastmoc) SPPS protocols. The crude cleaved peptides from each synthesis were compared by HPLC analysis. The overall aim of our work with Perloza is synthesis of resin-bound peptide ligands for affinity chromatography and antibody generation.(ABSTRACT TRUNCATED AT 250 WORDS)