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1.
Isoleucylcysteine dipeptide, a first intermediate peptide in bacitracin biosynthesis, was liberated from the enzyme protein and oxidized with manganese dioxide in dimethylsulfoxide. The resulting oxidation product was identified by thin-layer chromatography as 2-(2-methyl-l-oxobutyl)-thiazole-4-carboxylic acid which has been isolated from the hydrolysate of bacitracin F. This result shows that the intermediate dipeptide contains a thiazoline ring, and that the thiazoline ring is synthesized at the dipeptide stage in the process of peptide chain elongation in bacitracin biosynthesis. Improbability of non-enzymatic dehydrative cyclization of the dipeptide is discussed. 相似文献
2.
We describe a hidden Markov model, HMMSTR, for general protein sequence based on the I-sites library of sequence-structure motifs. Unlike the linear hidden Markov models used to model individual protein families, HMMSTR has a highly branched topology and captures recurrent local features of protein sequences and structures that transcend protein family boundaries. The model extends the I-sites library by describing the adjacencies of different sequence-structure motifs as observed in the protein database and, by representing overlapping motifs in a much more compact form, achieves a great reduction in parameters. The HMM attributes a considerably higher probability to coding sequence than does an equivalent dipeptide model, predicts secondary structure with an accuracy of 74.3 %, backbone torsion angles better than any previously reported method and the structural context of beta strands and turns with an accuracy that should be useful for tertiary structure prediction. 相似文献
3.
Experimental pulmonary cavity formation by mycobacterial components and synthetic adjuvants 总被引:4,自引:0,他引:4
An investigation was undertaken to determine the components of mycobacteria responsible for pulmonary cavity formation in tuberculosis. Rabbits received an intrapulmonary injection through the chest wall, of mycobacterial protein, II-p, mixed with either mycobacterial lipids, synthetic adjuvants or Nocardia cell wall skeleton as adjuvant. Six weeks later, they were killed and the lung lesions were examined. Cavities and necrosis were produced by the injection of II-p mixed with cord factor, Nocardia cell wall skeleton or N-acetylmuramyl dipeptide conjugated with long-chain branched fatty acids. Cavities were not produced by either the injection of II-p together with phospholipid, N-acetylmuramyl dipeptide (MDP), MDP-derivatives having no long-chain branched fatty acid, or by the injection of individual components of the mixtures. The results suggest that in tuberculosis a mycobacterial component with a long-chain branched fatty acid such as mycolic acid plays an important role in pulmonary cavity formation by enhancing the antigenicity of mycobacterial protein and helping it induce cell-mediated immunity at the site of the lesion. Passive transfer with sera from rabbits highly sensitized with tubercle bacilli failed to enhance cavity formation in the recipient animals. 相似文献
4.
R Shoeman T Coleman B Redfield R C Greene A A Smith I Saint-Girons N Brot H Weissbach 《Biochemical and biophysical research communications》1985,133(2):731-739
The regulation of the expression of three Escherichia coli met genes, metB, which codes for cystathionine gamma-synthetase (EC 4.2.99.9), metL, which codes for aspartokinase II-homoserine dehydrogenase II (EC 2.7.2.4-EC 1.1.1.3) and metJ, which codes for the methionine regulon aporepressor, has been studied using highly purified DNA-directed in vitro protein synthesis systems. In a system where the entire gene product is synthesized, the expression of the metB and metL genes is specifically inhibited by MetJ protein (repressor protein) and S-adenosylmethionine (AdoMet). In a simplified system that measures the formation of the first dipeptide of the gene product (fMet-Ala for the metJ gene), MetJ protein and AdoMet partially repress (approximately 40-60%) metJ gene expression. Thus, the metJ gene can be partially autoregulated by its gene product. 相似文献
5.
A protein sequence database (PFDB) containing about 11,000 entries is available for Macintosh computers. The PFDB can be easily updated by importing sequences from the PIR collection through the internet. The most important feature of the database is its organization in families of closely related sequences, each family being characterized by its average dipeptide composition [Petrilli (1993), Comput. Appl. Biosci.
2, 89–93]. This allows one to perform a rapid and sensitive protein similarity search by comparing the precalculated family dipeptide composition with that of the query sequence by a linear correlation coefficient. An example of an application in which a new protein was classsified by using a sequence of a fragment just 19 residues long is reported. 相似文献
6.
Deana C. Larkin Susan A. Martinis Deborah J. Roberts George E. Fox 《Origins of life and evolution of the biosphere》2001,31(6):511-526
The historical origin of the translation machinery remainsunresolved. Although the large 23S ribosomal RNA (rRNA) is almost certainly the catalytic component of the peptidyl transferase center in the modern ribosome, it is likely thatgreatly simplified systems were initially employed in the latestages of the prebiotic world. In particular, it has been suggested that small RNAs carrying amino acids were important forthe genesis of protein synthesis. Consistent with this, adipeptide, Ala-His, was previously claimed to be a prebioticallyfeasible catalyst mediating peptide bond formation in thepresence of aminoacylated tRNA and cognate mRNA template, in theabsence of other ribosomal components (Shimizu, 1996). We hereinreport a detailed study of putative dipeptide formation by Ala-His and RNAs carrying leucine. Based on the results presentedhere, it is unlikely that the dipeptide, Ala-His, catalyzessignificant levels of Leu-Leu dipeptide formation in solution. Aproduct is produced which can be readily mistaken for a dipeptidein the TLC separation systems employed in earlier work. We offerexplanations for the formation of this product as well as anotherunexpected product. The results presented here are consistentwith the notion that the translation machinery was likely basedon catalytic RNA from its very inception. 相似文献
7.
The branched polypeptide multi-L-arginyl-poly-L-aspartic acid, also called cyanophycin, is a water-insoluble reserve material of cyanobacteria. The polymer is degraded by a specific hydrolytic enzyme called cyanophycinase. By heterologous expression in Escherichia coli, a gene encoding cyanophycinase has been identified in the sequenced genome of Synechocystis sp. PCC 6803. The gene, designated cphB, codes for a protein of 29.4 kDa. The high level of expression of active cyanophycinase in E. coli from the Synechocystis gene allowed for its purification to electrophoretic homogeneity. The enzyme, which appears to be specific for cyanophycin, hydrolysed the polymer to a dipeptide consisting of aspartic acid and arginine. Based on inhibitor sensitivity and primary sequence, cyanophycinase appears to be a serine-type exopeptidase related to dipeptidase E [Conlin, C.A., Haakensson, K., Liljas, A. & Miller, C.G. (1994) J. Bacteriol. 176, 166-172]. 相似文献
8.
9.
Six different molecular dynamics simulations of Cyclophilin A, three with the protein free in water and three with the Ala-Pro
dipeptide bound to the protein, have been performed, and analysed with respect to structure and hydration of the active site.
The water structure in the binding pocket of the free Cyclophilin A was found to mimic the experimentally obtained binding
cis conformation of the dipeptide. Cyclophilin A is a peptidyl–prolyl cis–trans isomerase (PPIase), but the mechanism of the cis/trans isomerization is not exactly clear. This study was performed to understand better the binding between dipeptide and Cyclophilin
A, but also two previously proposed isomerization mechanisms are discussed.
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. 相似文献
10.
Sidorova MV Molokoedov AS Kudriavtseva EV Baldin MI Frid DA Ovchinnikov MV Bespalova ZhD Bushuev VN 《Bioorganicheskaia khimiia》2006,32(2):151-160
Two variants of the synthesis of tridecapeptide alloferon, the active principle of antiviral preparation allokine-alpha, were developed on the basis of fragment condensation in solution or on the Merrifield resin. The solid phase variant of the synthesis was shown to be more technological; it allows the preparation of the product at a higher total yield (40% vs. 17% for conventional synthesis in solution from the starting derivatives of the C-terminal dipeptide). The by-products formed during the synthesis of alloferon were identified. 相似文献
11.
12.
A plasmid-cloning vector system has been constructed which allows the production of fusion proteins with beta-galactosidase at the N terminus, followed by a recognition sequence for the site-specific protease, collagenase, and the foreign protein at the C terminus. A multicloning site allows the insertion of foreign genes in any translational reading frame. Fusion proteins were isolated by affinity chromatography on APTG-Sepharose. The foreign protein was released from the fusion product by collagenase cleavage. The vector was successfully utilized for the production of Escherichia coli single-stranded (ss) DNA-binding protein (SSB protein). The proteolytically released SSB protein resisted elution from an ss DNA-cellulose column with 1 M NaCl. 相似文献
13.
Crystal structure of the dipeptide binding protein from Escherichia coli involved in active transport and chemotaxis. 总被引:2,自引:1,他引:1
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P. Dunten S. L. Mowbray 《Protein science : a publication of the Protein Society》1995,4(11):2327-2334
The Escherichia coli periplasmic dipeptide binding protein functions in both peptide transport and taxis toward peptides. The structure of the dipeptide binding protein in complex with Gly-Leu (glycyl-L-leucine) has been determined at 3.2 A resolution. The binding site for dipeptides is designed to recognize the ligand's backbone while providing space to accommodate a variety of side chains. Some repositioning of protein side chains lining the binding site must occur when the dipeptide's second residue is larger than leucine. The protein's fold is very similar to that of the Salmonella typhimurium oligopeptide binding protein, and a comparison of the structures reveals the structural basis for the dipeptide binding protein's preference for shorter peptides. 相似文献
14.
To improve the pharmacological profile of tumor necrosis factor alpha (TNF-α), we have synthesized a new PEGylated prodrug,
PEG-vcTNF-α, using a cathepsin B-sensitive dipeptide (valine-citrulline, vc) to link branched PEG and TNF-α. PEG-modified
TNF-α without the dipeptide linker (PEG-TNF-α) and unconjugated TNF-α were also tested as controls. It was found for the first
time that TNF-α released from PEG-vcTNF-α was specifically dependent on the presence of cathepsin B. PEG-vcTNF-α induced higher
cytotoxicity and greater apoptosis against L929 murine fibrosarcoma cells than PEG-TNF-α. Reversal of these effects by a cathepsin-B
inhibitor confirmed that these effects were mediated by cathepsin B-specific release of TNF-α. In vivo pharmacokinetics studies demonstrated that the plasma stability of PEG-vcTNF-α was significantly increased compared to TNF-α.
Finally, the improved anticancer efficacy of PEG-vcTNF-α and the distinct activities among the three formulations confirmed
the positive contribution of both PEGylation and the dipeptide linkage to the improved drug-like properties of PEG-vcTNF-α.
The results here indicate that linking proteins and PEG via the cathepsin B-sensitive dipeptide may be a promising strategy
for developing protein therapeutics. 相似文献
15.
M. V. Sidorova A. S. Molokoedov E. V. Kudryavtseva M. I. Baldin D. A. Frid M. V. Ovchinnikov Zh. D. Bespalova V. N. Bushuev 《Russian Journal of Bioorganic Chemistry》2006,32(2):136-145
Two variants of the synthesis of tridecapeptide alloferon, the active principle of antiviral preparation allokine-alpha, were developed on the basis of fragment condensation in solution or on the Merrifield resin. The solid phase variant of the synthesis was shown to be more technological; it allows the preparation of the product at a higher total yield (40% vs. 17% for conventional synthesis in solution from the starting derivatives of the C-terminal dipeptide). The by-products formed during the synthesis of alloferon were identified. 相似文献
16.
Identification of a novel cleavage activity of the first papain-like proteinase domain encoded by open reading frame 1a of the coronavirus Avian infectious bronchitis virus and characterization of the cleavage products
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The coronavirus Avian infectious bronchitis virus (IBV) employs polyprotein processing as a strategy to express its gene products. Previously we identified the first cleavage event as proteolysis at the Gly(673)-Gly(674) dipeptide bond mediated by the first papain-like proteinase domain (PLPD-1) to release an 87-kDa mature protein. In this report, we demonstrate a novel cleavage activity of PLPD-1. Expression, deletion, and mutagenesis studies showed that the product encoded between nucleotides 2548 and 8865 was further cleaved by PLPD-1 at the Gly(2265)-Gly(2266) dipeptide bond to release an N-terminal 195-kDa and a C-terminal 41-kDa cleavage product. Characterization of the cleavage activity revealed that the proteinase is active on this scissile bond when expressed in vitro in rabbit reticulocyte lysates and can act on the same substrate in trans when expressed in intact cells. Both the N- and C-terminal cleavage products were detected in virus-infected cells and were found to be physically associated. Glycosidase digestion and site-directed mutagenesis studies of the 41-kDa protein demonstrated that it is modified by N-linked glycosylation at the Asn(2313) residue encoded by nucleotides 7465 to 7467. By using a region-specific antiserum raised against the IBV sequence encoded by nucleotides 8865 to 9786, we also demonstrated that a 33-kDa protein, representing the 3C-like proteinase (3CLP), was specifically immunoprecipitated from the virus-infected cells. Site-directed mutagenesis and expression studies showed that a previously predicted cleavage site (Q(2583)-G(2584)) located within the 41-kDa protein-encoding region was not utilized by 3CLP, supporting the conclusion that the 41-kDa protein is a mature viral product. 相似文献
17.
In a continuing effort to unravel the structural basis for isoform-selective inhibition of nitric oxide synthase (NOS) by various inhibitors, we have determined the crystal structures of the nNOS and eNOS heme domain bound with two D-nitroarginine-containing dipeptide inhibitors, D-Lys-D-Arg(NO)2-NH(2) and D-Phe-D-Arg(NO)2-NH(2). These two dipeptide inhibitors exhibit similar binding modes in the two constitutive NOS isozymes, which is consistent with the similar binding affinities for the two isoforms as determined by K(i) measurements. The D-nitroarginine-containing dipeptide inhibitors are not distinguished by the amino acid difference between nNOS and eNOS (Asp 597 and Asn 368, respectively) which is key in controlling isoform selection for nNOS over eNOS observed for the L-nitroarginine-containing dipeptide inhibitors reported previously [Flinspach, M., et al. (2004) Nat. Struct. Mol. Biol. 11, 54-59]. The lack of a free alpha-amino group on the D-nitroarginine moiety makes the dipeptide inhibitor steer away from the amino acid binding pocket near the active site. This allows the inhibitor to extend into the solvent-accessible channel farther away from the active site, which enables the inhibitors to explore new isoform-specific enzyme-inhibitor interactions. This might be the structural basis for why these D-nitroarginine-containing inhibitors are selective for nNOS (or eNOS) over iNOS. 相似文献
18.
Summary A new method of synthesis of Multiple Antigen Peptide System, in which the carrier is a core matrix with branched lysine and -alanine residues, is described. The antigenic peptide is separately synthesized in a protected form and coupled to the core with diisopropylcarbodiimide (DIPCDI) in presence of 1-hydroxy-benzotriazole (HOBt). This procedure has two major advantages: firstly, it allows an independent characterization (and purification) of the core and of the peptides; secondly, it allows a possible further coupling to a protein carrier, after an intermediate addition of Cys to the N-terminal amino acid. 相似文献
19.
Thomas Duerfahrt Sascha Doekel Theo Sonke Peter J L M Quaedflieg Mohamed A Marahiel 《European journal of biochemistry》2003,270(22):4555-4563
Microorganisms produce a large number of pharmacologically and biotechnologically important peptides by using nonribosomal peptide synthetases (NRPSs). Due to their modular arrangement and their domain organization NRPSs are particularly suitable for engineering recombinant proteins for the production of novel peptides with interesting properties. In order to compare different strategies of domain assembling and module fusions we focused on the selective construction of a set of peptide synthetases that catalyze the formation of the dipeptide alpha-l-aspartyl-l-phenylalanine (Asp-Phe), the precursor of the high-intensity sweetener alpha-l-aspartyl-l-phenylalanine methyl ester (aspartame). The de novo design of six different Asp-Phe synthetases was achieved by fusion of Asp and Phe activating modules comprising adenylation, peptidyl carrier protein and condensation domains. Product release was ensured by a C-terminally fused thioesterase domains and quantified by HPLC/MS analysis. Significant differences of enzyme activity caused by the fusion strategies were observed. Two forms of the Asp-Phe dipeptide were detected, the expected alpha-Asp-Phe and the by-product beta-Asp-Phe. Dependent on the turnover rates ranging from 0.01-0.7 min-1, the amount of alpha-Asp-Phe was between 75 and 100% of overall product, indicating a direct correlation between the turnover numbers and the ratios of alpha-Asp-Phe to beta-Asp-Phe. Taken together these results provide useful guidelines for the rational construction of hybrid peptide synthetases. 相似文献
20.
L Arfaoui-Aboulhab M Jouy S Fermandjian O Belhadj 《Archives de l'Institut Pasteur de Tunis》1986,63(2-3):289-298
The phenylalanyl circular dichroism (CD) bands of peptides were used to assay peptidase activity of carboxypeptidase A (EC.3.4.12.2.). Hippuryl-L-phenylalanine has a sharp, negative CD band at 254 nm whilst L-phenylalanine (the optically active product) has positive CD. Thus the hydrolysis of this substrate as well as the inhibition effect of dipeptides, may be measured from the CD change at 254 nm. The addition of the dipeptide GLy-Tyr to carboxypeptidase A makes the CD spectrum more positive in the region from 270-295 nm. This alteration can result from the tyrosyl and tryptophanyl CD bands of the protein as well as from the tyrosyl CD band of the inhibitor. 相似文献