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T L Voiushina

Publications and source records attributed to T L Voiushina.

14 recordsLinked to original sources
PubMed2004

[A new method for determination of antithrombin III and its diagnostic significance].

A new assay based on using an original chromogenic substrate (z-Ala-Ala-Arg-pNa) and designed for the determination of antithrombin III is described in the paper. It was used in examinations of patients with malignancy after surgery. The content of antithrombin III was detected to be lower in 12% of cases in 2 days after surgery. Additionally, patients with DIC, as complications after surgery, were examined to find out that the antithrombin III content was decreasing in them.

Adolescent↗
PubMed2000

[A method for determining plasminogen with a Russian chromogenic substrate and its diagnostic significance].

Measurement of plasminogen, the key component of fibrinolysis system, is one of the basic methods for estimation of fibrinolysis. Methods based on the use of chromogenic substrates are often used in diagnosis. Plasminogen measurements are important for laboratory diagnosis of thrombophilia caused by deficiency or abnormalities of this fiber, for detection and evaluation of the DIC syndrome, and for monitoring the treatment by fibrinolytic preparations (streptokinase, t-PA, urokinase, etc.). An original chromogenic substrate having no foreign analogs has been created at Institute of Genetics and Selection of Industrial Microorganisms and Research Center of Hematology (Moscow). Unlike previously described plasmin substrates, pNa has been obtained by microbiological methods with Russian commercial enzymes subtilisine 72 and megaterine. This paper presents the results of plasminogen measurements in patients with DIC with the use of the original chromogenic substrate. The results were compared with those of tests with Berihrom-Plasminogen diagnostic kit (Behringwerke AG).

Adolescent↗
PubMed1997

[Affinity sorbents with tripeptide morpholine ligands for isolation of proteases].

New affine sorbents were synthesized involving tripeptide morpholides H-Ala-Ala-Leu-Mrp and H-D-Ala-Leu-Arg-Mrp as ligands that mimic substrates of subtilisin-like proteases and kallikrein, respectively. These were used for the isolation and purification of several proteases: trypsin, pepsin, alpha-chymotrypsin, thrombin, kallikrein, and termitase and were also efficient in the isolation of proteolytic enzymes from complex mixtures, such as the urine of children suffering from glomerulonephritis, hepatopancreas of Kamchatka crab, and dandelion roots. The ligands are competitive inhibitors of a number of proteases, and therefore, they were supposed to interact with the substrate binding sites in these enzymes.

Animals↗
PubMed1996

[Proteolytic enzymes from Streptomyces fradiae: a metalloendopeptidase, subtilisin-like, and trypsin-like proteinases].

Three proteolytic enzymes-the metalloproteinase, SFMP, and two serine proteinases, SFSP and SFTP-have been isolated and purified from the culture fluid of Streptomyces fradiae using chromatography on bacitracin-silochrome, bacitracin-Sepharose, DEAE-cellulose and fractionation by ammonium sulfate. Study of physico-chemical and functional properties of the enzymes and structural analysis revealed that SFMP is a cysteine-containing metalloendopeptidase with M(r) of 36 kDa, has a peak activity for synthetic substrates at pH 7.0-7.5 and at 60-65 degrees C and is stable at pH 7.0-9.0. The serine proteinase SFSP is related to subtilisin-like enzymes, has a M(r) of 29 kDa and a pH optimum at 7.5-8.5 at temperature up to 50 degrees C. The proteinase is stable at pH 4.0-9.0 and retains 30% of its activity at 70 degrees C. The other serine proteinase, SFTP, has a M(r) of 26 kDa and is related to trypsin-like enzymes. Its activity for synthetic substrates of trypsin is maximal at pH 6.8-8.8 at 50 degrees C. The enzyme is stable at pH 4.5-8.5 and at temperature below 50 degrees C. It has been shown that Streptomyces fradiae, like Streptomyces griseus and other Streptomycetes, possesses an ability to secrete serine proteinases (SFSP and SFTP) related to two evolutionally distinct families of serine proteinases, i.e., subtilisin and chymotrypsin families. SFMP and SFSP have been isolated and characterized for the first time.

Amino Acid Sequence↗
PubMed1991

[Enzymatic synthesis of acyl peptides containing p-nitroanilides of basic amino acids].

A method is suggested for synthesis of acylpeptides, containing arginine or lysine p-nitroanilides at the C-terminus, via the acyl transfer reaction catalyzed by the Bacillus subtilis serine proteinase. Acyl-di- and acyltripeptide ethers with L- and D-amino acids were used as the carboxyl component taken in a twofold excess. When the concentration of dimethylformamide increases, the hydrolysis of the initial ether and the reaction product diminishes. Because of the enzyme inactivation by dimethylformamide the latter's optimal concentration is 70-80%.

Acylation↗
PubMed1991

[Isolation and properties of serine proteinase from Aspergillus oryzae].

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.

Amino Acid Sequence↗
PubMed1991

[Substrate specificity of the serine proteinase from Bacillus subtilis, strain 72].

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.

Amino Acids↗
PubMed1987

[Synthesis of p-nitroanilides of acylated peptides catalyzed by thermolysin].

Thermolysin-catalysed synthesis of p-nitroanilides of acylpeptides of general formula Z-A1-A2-pNA (A1 = Thr, Ala, Val, Leu; A2 = Leu, Phe) and stepwise synthesis of p-nitroanilides of acyltetrapeptides of general formula Z-A1-A2-A3-A4-pNA (A1, A2 = Gly,Ala; A3, A4 = Ala, Leu, Phe) from Z-A1-A2-OH and A3-pNA and then from Z-A1-A2-A3-OH and A4-pNA have been carried out; pNA group was eliminated enzymatically. Increase in solubility of the product in the reaction mixture diminishes its yield. Minimal amount of thermolysin providing a substantial yield of reaction product depends on structure of both amino and carboxylic components. In many cases the molar ratio of the enzyme and starting substances could be decreased to 1:10(6) as compared with the generally used ration 1:10(3)-1:10(4).

Amino Acids↗
PubMed1978

[Inhibitors of exonuclease A5].

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.

Cytidine Diphosphate↗
PubMedDate not supplied

[Synthesis of modified peptides with C-terminal alpha-amino aldehydes].

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.

Aldehydes↗
PubMedDate not supplied

[Glutamyl endopeptidase. Structure, function, practical use].

Special features of the structural organization of serine proteases belonging to a new subfamily of glutamyl-specific endopeptidases, which possess an extremely strict substrate specificity, are discussed. Some areas of the practical application of these enzymes are considered. The English version of the paper: Russian Journal of Bioorganic Chemistry, 2003, vol. 29, no. 6; see also http://www.maik.ru.

Models, Molecular↗
PubMedDate not supplied

[The interaction of carriers with proteolytic enzymes used for enzymatic synthesis of peptides in organic solvents].

The adsorption of some proteolytic enzymes on inorganic solvents (Silochromes C-80 and C-120, macroporous glass CPG-10, and celite 535) and the effect of sorption-desorption processes on the activity and stability of the enzymes have been studied. The ability of the enzymes to be adsorbed on the carrier depended on the specific surface of the carrier and properties of the enzyme. Adsorption-desorption processes did not affect the activity of the enzymes. Acetonitrile had no noticeable effect on the activity of the enzyme adsorbed on the inorganic carrier. The enzymes adsorbed on such carriers catalysed reactions in media with a low (4-5%) content of water, since the carrier seems to protect the enzyme from inactivation with organic solvents. The loading of carrier with enzyme influenced the rate of the enzymatic reaction, the optimal loading corresponding the value of the maximum adsorption of the enzyme on the carrier.

Adsorption↗
PubMedDate not supplied

[Activity of endogenous proteinases in glomerulonephritis in children].

Protease activities in serum and urine of 116 children with glomerulonephritis and 16 healthy children were tested using chromogenic peptide substrates Z-Dala_Leu-Arg-pNA and Glp-Ala-ALa-Leu-pNa. We found the dependence between activity of serine proteinases and clinical, morphological forms of primary glomerulonephritis and hypertension.

Adolescent↗