[A novel endogeneous inhibitor from hepatopancreas of Kamchatka crab (Paralithodes camtschaticus)].

A novel endogeneous inhibitor from hepatopancreas of Kamchatka crab (Paralithosed camtschaticus) was isolatyed. The inhibitor was purifeid through fractional affinity chromatography on gramicidin-diasorb followed by gel-filtration at Sephadex G-100. The inhibitor PC is a protein (M, 66 kDa) and active against serine collagenolytic protease PC at temperature optimum 15-20 degrees C, stable at 4-40 degrees C and was completely inactivated after heating to 50 degrees C and higher. 0.9-20% NaCl is necessary for its inhibitor activity. The inhibitor was found to slow down cell spreading in vitro in cell type-dependent manner. Fibroblasts are most prone to inhibitory effect, epithelial tumor derived cells show medium susceptibility, while fibrosarcoma cells were not affected.

[Anticoagulative and anticomplementary activity of endogenous inhibitor preparation from hepatopancreas of red king crab (Paralithosed camtschaticus) towards human blood].

Serpins (SERine Protease INhibitors)--is large and diverse group of proteins with similar structures, which can inhibit both serine and cysteine proteases by an irreversible suicide mechanism. A novel serpin from hepatopancreas of Red King Crab (Paralithosed camtschaticus) was obtained and was studied its effect on the process of human blood plasma clotting. The investigated serpin shows a noticeable anticoagulative activity, which increases dramatically in the combined action with heparine. Though the inhibitor has almost no effect on thrombin, it inhibits C1s (C1-esterase). We studied the action of the serpin from P. camtschaticus on C1s via its competitive inhibition by C1 inhibitor and the novel enzyme. The calculated inhibition constant of the serpin from P. camtschaticus towards C1s is 2.02 +/- 0.71 M. Unlike C1 inhibitor, the novel serpin from P. camtschaticus doesn't suppress fibrinolysis and at the same time prevents blood clotting. These features may be of interest for medical purposes.

[Endocellular aminopeptidase from Astasia longa].

A new aminopeptidase was isolated from the biomass of the flagellate Astasia longa by precipitation with ammonium sulfate, gel filtration, and affinity chromatography on Arginine-Silochrome in 41% yield and with purification degree 490. The enzyme is irreversible inhibited by mercury chloride, EDTA, o-phenanthroline and, partially, bestatin and zinc chloride. It has an optimum pH 8.5 toward the hydrolysis of a synthetic chromogenic substrate Ala-pNA. The enzyme molecular mass is 45 kDa, isoelectric point 5.5, and temperature optimum 45 degrees C. The enzyme most effectively hydrolyzes p-nitroanilides of alanine, arginine, and leucine; it is classified as metalloaminopeptidase.

[Design of a recombinant antigen for the diagnosis and prevention of larval hydatidosis].

The paper deals with the design of recombinant vector make-ups, with the expression of Echinococcus hybrid proteins, and with the study of their immunogenic properties. Theoretical rationale is given for the choice of the parasitic gene superexpression system (E. coli cells of SG 13009 strain--recombinant plasmid pQE/EgF). The authors show that the use ofpolymerase chain reaction with oligonucleotide primers homologous to the structures of unique genes coding for Echinococcus antigen is promising for the run of the preparative quantities of fragments of these genes. They consider the basic stages of obtaining hybrid EgF antigen: the isolation of genomic DNA from Echinococcus protoscolexes; the run of preparative quantities of an EgF DNA fragment; the obtaining of vector pQE plasmid DNA; the design of a recombinant make-up; the screening of positive clones; the recombinant plasmid expression of hybrid protein and its purification. The commission tests of EgF antigen in enzyme immunoassay using 93 human serum samples revealed the following: the sensitivity and specificity were 83.8 and 77.4%, respectively. The recombinant protein of EgF was found to exert a significant protective action on the development of E. multilocularis larvocysts in non-inbred albino mice.

[Trypsin inhibitor from Amaranth (Amaranthys cruentus) leaves]. / Ingibitor tripsina iz list'ev amaranta (Amaranthus Cruentus)

A protein that inhibited the proteolytic activity of trypsin was isolated from amaranth leaves (Amaranthus cruentus) by affinity chromatography on trypsin-Sepharose. The inhibition was noncompetitive (with n-nitroanilide-N-alpha-benzoyl-DL-arginine as substrate) and had a Ki of 11.87 x 10(-7) 7 M. The protein caused a weaker inhibitory effect on chymotrypsin, had no effect on subtilisin, displayed a molecular weight of 8 kDa, and contained no cysteine residues.