The segment condensation of peptides on a solid phase (Aminosilochrom) in organic medium catalyzed by a subtilisin complex with sodium dodecylsulfate was studied. The dependence of the efficiency of the enzymatic coupling of tripeptides with the basic structure X-Ala-Ala-Y-OMe [where X = Z, Boc, or Dnp and Y = Leu or Glu(OMe)] on the spacer content on the support and on the structure of the acylating component was investigated. The tripeptide segments were successively coupled to Aminosilochrom containing the Met-Ala-Gly spacer, and the peptidylaminosilochroms Dnp-Ala-Ala-Leu-Ala-Ala-Leu-Ala-Ala-Glu(OMe)-Met-Ala-Gly-A and Dnp-Ala-Ala-Leu-Ala-Ala-Glu(OMe)-Ala-Ala-Leu-Met-Ala-Gly-A (A is the Aminosilochrom residue) were obtained in satisfactory yields. It was shown by these examples that the second and third segments are attached in yields higher than that for the first segment and the coupling efficiency does not depend on the amino acid composition of the acylating component.

Amino Acids↗

PubMedDate not supplied

Proteinase-catalyzed peptide synthesis in concentrated solutions of urea and other denaturing agents.

Pepsin successfully catalyzed the synthesis of several hydrophobic octa- and decapeptides in dimethylformamide-water solutions containing concentrated urea at pH 4.65. The factors that influence peptide synthesis in the presence of urea were studied using condensation of the tripeptides Z-Ala-Ala-Phe-OH and H-Leu-Ala-Ala-OCH3 as a model. The dependence of Z-Ala-Ala-Phe-Leu-Ala-Ala-OCH3 yield on pepsin concentration and pH, as well as the behavior of pepsin during peptide synthesis were studied. It was shown that pepsin catalyzed the synthesis of Z-Ala-Ala-Phe-Leu-Ala-Ala-OCH3 in guanidine hydrochloride and sodium dodecyl sulfate solutions. Other proteinases, subtilisin and thermolysin, were applied for the synthesis of p-nitroanilides of tri- and tetrapeptides in urea solutions. Proteinase-catalyzed peptide synthesis in the presence of denaturing agents might help to overcome the limitations caused by poor solubility of the starting peptide derivatives, although this effect is sometimes counterbalanced by the product solubility.

Amino Acid Sequence↗

PubMedDate not supplied

[Preparation of highly puried pepsin by aminosilichrome chromatography].

The procedure for preparing highly purfied pepsin by chromatography on silicon dioxide with attached aminogroups (aminosilochrome) has been devised. Pepsin is eluated step=by-step: 0.0025 M and 0.05 M HCl. Results of disc-electrophoresis of purified pepsin bive evidence that the resultant preparation contains no admixtures. The activity of purified pepsin is 69 units per mg.

Chromatography, Affinity↗

PubMedDate not supplied

[Production of proteinases in the process of developing cultures of Streptomyces thermovulgaris T-54].

The secretion of some proteolytic enzymes by Streptomyces thermovulgaris T 54 was studied using artificial chromogenic substrates of proteinases. Maximum accumulation of the enzymes hydrolysing Z-Glu-pNA and Z-Ala-Ala-Leu-pNA occurred during autolysis of the culture. Two peaks of activity were observed, when DNP-Gly-Gly-Ile-Arg was used as a substrate, one of them being correspondent to prevalence of dense colonies in the culture and the other to the prevalence of friable networks of hyphae.

Amino Acid Sequence↗

PubMedDate not supplied

[The effect of Legionella pneumophila metalloproteinase on certain human blood proteins. Cleavage of anti(1)-antitrypsin and acid-stable serine proteinase inhibitors].

Antitryptic activity of human blood serum was decreased after incubation with metalloproteinase from Legionella pneumophila. The enzymatic activity depends on the time of incubation as well as on the ratio between the enzyme content and blood serum total protein. Cross immunoelectrophoresis, involving monospecific rabbit antiserum towards the alpha 1-antitrypsin, demonstrated highly effective hydrolysis of alpha 1-antitrypsin by the metalloproteinase. As shown by polyacrylamide gel electrophoresis the metalloproteinase hydrolyzed acid stable inhibitor of serine proteinases into fragments with distinct loss of the inhibitor activity. Thermolysine hydrolyzed similarly the proteins studied but at a lower rate. The metalloproteinase from L. pneumophila appears to be mainly responsible for production of utilizable components from protein substrates involved in vital activity of the bacteria. It may not be excluded that the enzyme is able to impair some host protective mechanisms.

Electrophoresis, Polyacrylamide Gel↗

PubMedDate not supplied

[The properties of HIV protease synthesized in E. coli cells].

A hybrid plasmid pPR6 was constructed containing BgII-EcoRI fragment of the pol region of HIV (strain IIIB) genome which determined the synthesis of virus-specific protease. Extracts of E. coli DN5/pPR6 bacteria provided for specific hydrolysis of hybrid protein p165 (the N-terminus of which is presented by complete beta-galactosidase and the C-terminus by duplicated area of virus-specific precursor p55 containing a site for virus-specific protease located at the border of proteins p17 and p24) with formation of products having molecular weights of 19, 42, 28, 23, and 19 kD. Polypeptides 119K, 23K, and 19K are products of complete hydrolysis, and 42K a result of partial cleavage. The kinetics of hydrolysis in relation to pH values of the reaction mixture was analysed. It is suggested that the reported system of HIV protease activity determination be used for screening of potential inhibitors of this enzyme.

Electrophoresis, Polyacrylamide Gel↗

PubMedDate not supplied

[Determination of aspartame--methyl ester of L-aspartyl-L-phenyl- alanine using the amino acid analyzer].

A technique for quantitative determining of the synthetic sweetener aspartame. NH2-Asp-Phe-OMe and its hydrolysis product, NH2-Asp-Phe-OH, is proposed based on the use of an amino acid analyzer.

Amino Acids↗

PubMedDate not supplied