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1.
Zh V Potetinova E I Mil'gotina V A Makarov T L Voiushina 《Bioorganicheskaia khimiia》2001,27(3):163-173
Various synthetic approaches to modified peptides with the C-terminal aldehyde group, capable of inhibiting a number of proteolytic enzymes belonging to the classes of thiol, serine, and aspartyl proteases, are considered. Both chemical methods, including solid phase peptide synthesis now widely used, and biocatalytic synthetic methods for obtaining these substances are discussed in detail. 相似文献
2.
T I Vagonova N M Ivanova O M Khodova T L Voiushina V M Stepanov 《Biokhimii?a (Moscow, Russia)》1991,56(1):125-135
A serine proteinase having an activity optimum at pH 6.7-8.2 has been isolated from amylorisine P-10x (a mixture of Aspergillus oryzae enzymes) by chromatography on DEAE-Sephadex A-50 and bacitracin Sepharose 4B. The proteinase is fully inactivated by phenylmethylsulfonylfluoride and diisopropylfluorophosphonate, the specific inhibitors of the enzyme, and has a pI at pH 7.5. The molecular mass of serine proteinase is 30000 Da; its amino acid composition appears as: Met2, Asp33, Thr18, Ser29, Glu21, Pro9, Glu32, Ala38, Val24, Ile16, Leu15, Tyr8, Phe8, His8, Lys18, Arg4, Trp6. The N-terminal sequence of the serine proteinase: Gly-Leu-Thr-Thr-Gln-Lys-Ser-Ala-Pro-Trp-Gly-Leu-Gly-Ser-Ile-Ser-Xaa-Lys- Gly-Gln-Gln-Ser-Thr-Asp-Tyr-Ile-Tyr, which coincides practically completely with the corresponding sequence of alkaline proteinase of A. oryzae, ATCC20386, has been determined. Similar to subtilisin, the enzyme catalyzes the condensation of leucine and alanine p-nitroanilides with N-benzyloxycarbonyl-alanyl-alanine and glycyl-alanine methyl esters. 相似文献
3.
Background
Protein translocation across the membrane of the Endoplasmic Reticulum (ER) is the first step in the biogenesis of secretory and membrane proteins. Proteins enter the ER by the Sec61 translocon, a proteinaceous channel composed of three subunits, α, β and γ. While it is known that Sec61α forms the actual channel, the function of the other two subunits remains to be characterized.Results
In the present study we have investigated the function of Sec61β in Drosophila melanogaster. We describe its role in the plasma membrane traffic of Gurken, the ligand for the Epidermal Growth Factor (EGF) receptor in the oocyte. Germline clones of the mutant allele of Sec61β show normal translocation of Gurken into the ER and transport to the Golgi complex, but further traffic to the plasma membrane is impeded. The defect in plasma membrane traffic due to absence of Sec61β is specific for Gurken and is not due to a general trafficking defect.Conclusion
Based on our study we conclude that Sec61β, which is part of the ER protein translocation channel affects a post-ER step during Gurken trafficking to the plasma membrane. We propose an additional role of Sec61β beyond protein translocation into the ER. 相似文献4.
Sequence of a cDNA for mouse thymidylate synthase reveals striking similarity with the prokaryotic enzyme 总被引:11,自引:0,他引:11
Perryman SM; Rossana C; Deng TL; Vanin EF; Johnson LF 《Molecular biology and evolution》1986,3(4):313-321
We report the nucleotide sequence of a cloned cDNA, pMTS-3, that contains a
1-kb insert corresponding to mouse thymidylate synthase (E.C. 2.1.1.45).
The open reading frame of 921 nucleotides from the first AUG to the
termination codon specifies a protein with a molecular mass of 34,962
daltons. The predicted amino acid sequence is 90% identical with that of
the human enzyme. The mouse sequence also has an extremely high degree of
similarity (as much as 55% identity) with prokaryotic thymidylate synthase
sequences, indicating that thymidylate synthase is among the most highly
conserved proteins studied to date. The similarity is especially pronounced
(as much as 80% identity) in the 44-amino-acid region encompassing the
binding site for deoxyuridylic acid. The cDNA sequence also suggests that
mouse thymidylate synthase mRNA lacks a 3' untranslated region, since the
termination codon, UAA, is followed immediately by a poly(A) segment.
相似文献
5.
M Iu Gololobov I P Morozova T L Voiushina E A Timokhina V M Stepanov 《Biokhimii?a (Moscow, Russia)》1991,56(2):230-240
A comparative study of the hydrolysis of various p-nitroanilide substrates (Z-A2-A1-pNA, Z-A3-A2-A1-pNA, and Z-A4-A3-A2-A1-pNA, where A1-An are various amino acid residues, Z is the benzoyloxycarbonylic group and pNA is the p-nitroanilide group), catalyzed by serine proteinase from Bacillus subtilis strain 72, was carried out. It was found that depending on the substrate structure, the hydrolysis may involve both the peptide-p-nitroaniline and the amino acid-amino acid bonds. A kinetic analysis of substrate hydrolysis occurring simultaneously at these two bonds was carried out. The physico-chemical meaning of the kinetic parameters of the given scheme was determined. The quantitative estimation of the enzyme specificity with respect to both hydrolyzing bonds can be found by using the parameters calculated during the analysis of the kinetic curve of p-nitroaniline production. It was found that according to their specificity the amino acid residues at position A1 can be arranged in the following order: L-Leu greater than P-Phe greater than L-Ile greater than L-Ala. The beta-branched amino acid residues, L-Val and L-Ile, do not bind to subsite S1. If these residues occupy position A1, the substrate splitting occurs exclusively between residues A1 and A2. The tetrapeptide N-protected p-nitroanilide substrates are also hydrolyzed at this bond. Partial hydrolysis of the amino acid-amino acid bond between residues A1 and A2 occurs in two cases: i) when residue A1 is loosely bound to subsite S1 and/or, ii) when residue A2 is firmly bound to subsite S1. 相似文献
6.
7.
T H L'vova R I Tatarskaia T L Voiushina S Z Simonian V R Varlamov D R Val'kovskii S V Rogozhin A A Baev 《Biokhimii?a (Moscow, Russia)》1978,43(2):350-364
Over 30 compounds resembling to or being structural elements of the minimal substrate of exonuclease A5 were tested for their ability to inhibit the reaction catalyzed by this nuclease. The compounds containing less than two phosphate groups were shown to possess a low inhibitory activity, if any. CDP, double-stranded DNA and nucleoside-3',5'-diphosphates (pNp) proved to be effective exonuclease A5 inhibitors. Pyrophosphate stimulated the reaction in the case of low molecular weight substrates only. For the inhibitory activity of pNp to occur, the intactness of the nucleoside moiety as a whole was shown to be necessary, the activity level depending on the structure of both the base and the sugar components. A competitive mechanism of the inhibitory action was demonstrated for pTp, pdCp, pdGp, pdAp and pUp and the Ki values were determined. The affinity for the inhibitor decreased in the following order: pdCp greater than or equal to pTp greater than pdGp greater than pdAp. Ki for pdCp and pTp were found to be approximately 3.10(-6) M. The investigation of the inhibition mechanism as well the determination of Ki were accomplished with the help of homogenous low molecular weight substrates--ApApA and the phosphoamide MeOPheNH(pdA)2. These were chosen after kinetic parameters determination of the hydrolysis of 22 exonuclease A5 substrates, predominatly of the RpNpN type. On the basis of data obtained the specificity of exonuclease A5 is also discussed. Possible usefulness of immobilized competitive inhibitors of the pNp type not only for single nuclease isolation but for the separation of a mixture of different nucleases is considered. This possibility is based on the almost universal inhibitory effect of pNp on different nucleases and at the same time on their different affinity for the enzymes. In particular, this approach might be useful for the elimination of exonucleases and some other nucleolytic enzymes from the preparations of endonucleases-restrictases. 相似文献
8.
Brigida TL Lucena Billy M dos Santos João LS Moreira Ana Paula B Moreira Alvaro C Nunes Vasco Azevedo Anderson Miyoshi Fabiano L Thompson Marcos Antonio de MoraisJunior 《BMC microbiology》2010,10(1):298
Background
Bacteria may compete with yeast for nutrients during bioethanol production process, potentially causing economic losses. This is the first study aiming at the quantification and identification of Lactic Acid Bacteria (LAB) present in the bioethanol industrial processes in different distilleries of Brazil. 相似文献9.
Role of the reticulum in the stability and shape of the isolated human erythrocyte membrane 总被引:5,自引:7,他引:5 下载免费PDF全文
In order to examine the widely held hypothesis that the reticulum of proteins which covers the cytoplamsic surface of the human erythrocyte membrane controls cell stability and shape, we have assessed some of its properties. The reticulum, freed of the bilayer by extraction with Triton X-100, was found to be mechanically stable at physiological ionic strength but physically unstable at low ionic strength. The reticulum broke down after a characteristic lag period which decreased 500-fold between 0 degrees and 37 degrees C. The release of polypeptide band 4.1 from the reticulum preceded that of spectrin and actin, suggesting that band 4.1 might stabilize the ensemble but is not essential to its integrity. The time-course of breakdown was similar for ghosts, the reticulum inside of ghosts, and the isolated reticulum. However, at very low ionic strength, the reticulum was less stable within the ghost than when free; at higher ionic strength, the reverse was true. Over a wide range of conditions the membrane broke down to vesicles just as the reticulum disintegrated, presumably because the bilayer was mechanically stabilized by this network. The volume of both ghosts and naked reticula varied inversely and reversibly with ionic strength. The volume of the naked reticulum varied far more widely than the ghost, suggesting that its deformation was normally limited by the less extensible bilayer. The contour of the isolated reticulum was discoid and often dimpled or indented, as visualized in the fluorescence microscope after labeling of the ghosts with fluoroscein isothiocyanate. Reticula derived from ghosts which had lost the ability to crenate in isotonic saline were shriveled, even though the bilayer was smooth and expanded. Conversly, ghosts crenated by dinitrophenol yielded smooth, expanded reticula. We conclude that the reticulum is a durable, flexible, and elastic network which assumes and stabilizes the contour of the membrane but is not responsible for its crenation. 相似文献
10.
Molecular evolution of a multigene family in group A streptococci 总被引:15,自引:0,他引:15
The emm genes are members of a gene family in group A streptococci (GAS)
that encode for antiphagocytic cell-surface proteins and/or
immunoglobulin-binding proteins. Previously sequenced genes in this family
have been named "emm," "fcrA," "enn," "arp," "protH," and "mrp"; herein
they will be referred to as the "emm gene family." The genes in the emm
family are located in a cluster occupying 3-6 kb between the genes mry and
scpA on the chromosome of Streptococcus pyogenes. Most GAS strains contain
one to three tandemly arranged copies of emm-family genes in the cluster,
but the alleles within the cluster vary among different strains.
Phylogenetic analysis of the conserved sequences at the 3' end of these
genes differentiates all known members of this family into four
evolutionarily distinct emm subfamilies. As a starting point to analyze how
the different subfamilies are related evolutionarily, the structure of the
emm chromosomal region was mapped in a number of diverse GAS strains by
using subfamily-specific primers in the polymerase chain reaction. Nine
distinct chromosomal patterns of the genes in the emm gene cluster were
found. These nine chromosomal patterns support a model for the evolution of
the emm gene family in which gene duplication followed by sequence
divergence resulted in the generation of four major-gene subfamilies in
this locus.
相似文献