Cloning and characterization of serpin from red king crab Paralithodes camtschaticus.

Serpins are a family of serine protease inhibitors that are involved in numerous physiological processes and are known to regulate innate immunity pathways. To advance our understanding of their role in P. camtschaticus, a commercially significant species, we cloned and characterized a serpin from this species, designated serpin PC, that has anticoagulant and anticomplement effects on human blood. We found that serpin PC is a secreted protein with a typical serpin-like primary structure that is similar to other known crustacean serpins. Recombinant serpin PC was found to have inhibitory activity against R/K-specific bovine cationic trypsin. The reaction proceeds through the formation of a stable covalent complex of peptidase with P1 residue R383 of serpin PC. This interaction is characterized by a relatively high overall inhibition constant kass=(2.3 ±â€¯0.7) × 106 M-1s-1 and an SI of 4.7 ±â€¯0.8. Protein localization by western blotting showed that serpin PC is present in the muscles and, to a lesser extent, the heart, whereas it is transcribed predominantly in hemocytes and the heart. Through peptidase activity profiling of hemocytes and plasma, we found that serpin PC inhibits at least two R/K-specific activities and showed that it inhibits phenoloxidase (PO) activity induction in hemocytes.

Preparation of Recombinant Serpin from Red King Crab Paralithodes сamtschaticus for Biomedical Research Purposes.

Genetic constructs with different leader sequences for intra- and extracellular expression of the target protein were generated and an original method for effective selection of clones with maximum expression was developed. For intracellular expression in the Pichia pastoris system, seprin content in cells was 6 mg/liter.

[Investigation of the effect of oil and flour from wheat germ meal on lipid metabolism of students and teachers of the university].

The results of investigation of alimentary correction of lipid metabolism under the admin­istration of processed products from wheat germ - oil (with the content of policosanol at least 1.5-8.0 mg/100 g, vitamin E - 180-200 mg/100 g, PUFA - 60-65%) and cake flour (with the content of protein - 30-35%, oil with analogue composition -5-7%, digestible carbohydrates - 45-47%, fiber - 18-26%, vitamins B1, B3, B6, B9, E, PP, minerals and trace elements - Zn, Mn, K, Fe, Se, P) are presented. Volunteers among teachers and students of the university aged 16 to 65 years daily consumed wheat germ oil obtained by cold pressing in an amount of 3.5 g, regardless of the meal within 30 days. Then a part of them (30 persons) consumed daily 50 g of oil cake obtained after pressing oil, which provided the intake of the same amount of oil (3.5 g). Lipid metabolism param­eters were monitored in experiment participants before receiving the processed products of wheat germ, after germ meal intake and beyond 30 and 60 days after consumption of wheat germ. Data analysis was carried out on three age groups: 16-24, 25-44 and 45- 65 years. All participants of the experiment showed a reduction in total cholesterol level by 6-8%, increasing the concentration of HDL cholesterol by 3-24%, lowering LDL cholesterol concentrations by 4-21%, reduction of triglyceride concentration by 12-24%, a positive correction of atherogenic factor values by for 10-25%. Prolonged action of the investigated foods was established: lipid metabolism parameters in the tested group were better than in the control group after 30 days of intake discontinuation of oil or wheat germ flour, the positive adjustment effect disappeared 60 days after consuming the products. The findings demonstrate a positive effect on the normalization of lipid metabolism when cake flour of wheat germ was administered in daily food ration, similar to the effect of oil intake, which is important for the prevention of cardiovascular diseases and atherosclerosis. Given the significant production of cake flour of wheat germ (up to 90-95% of the raw material) and its not high cost as a secondary biological resource, this product can be recommended to the introduction in the diet of organized groups, including socially vulnerable groups.

Simultaneous dual-wavelength stimulated Raman scattering in Ba(MoO4)x(WO4)(1-x) solid solution single crystals.

The simultaneous dual-wavelength stimulated Raman scattering (SRS) of the same order Stokes with 532 nm nanosecond laser pulse excitation was found for the first time to the best of our knowledge in Ba(MoO4)x(WO4)1-x single crystals. The results of studies of some peculiarities of this phenomenon were reported.

Active microwave pulse compressor using an electron-beam triggered switch.

A high-power active microwave pulse compressor is described that operates by modulating the quality factor of an energy storage cavity by means of mode conversion controlled by a triggered electron-beam discharge across a switch cavity. This Letter describes the principle of operation, the design of the switch cavity, the configuration used for the tests, and the experimental results. The pulse compressor produced output pulses with 140-165 MW peak power, record peak power gains of 16∶1-20∶1, and FWHM pulse duration of 16-20 ns at a frequency of 11.43 GHz.

[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.

[Pharmacogenomical aspects of gastroesophageal reflux disease].

The article covers the modem concept of gastroesophageal reflux disease (GERD) pharmacogenomics. Special attention is payed to polymorphism of CYP2C19 gene and its influence on the metabolism of modern antisecretory agents, such as proton pump inhibitors (PPI). The authors summarize data which demonstrate that the differences in the pharmacodynamics and clinical effects of PPI are caused by CYP2C19 gene polymorphism. The paper shows that this factor should be considered when planning GERD therapy with PPI in order to achieve a complete remission and make the therapy economically rational.

[Food additive (bifeinol, detovit, squalenol) in the treatment of patients with metabolic syndrome]. / Pishchevaia dobavka (bifeinol, detovit, skvalenol) v lechenii bol'nykh metabolicheskim sindromom.

For 2 months, 33 patients with the metabolic syndrome (MS) took a biologically active food additive as bifeinol, detovit, squalenol, which contained vitamins, minerals, essential amino acids, dietary cellulose, and a complex of polysaturated fatty acids. This yielded a positive result as decreases in body weight by 8%, in the levels of cholesterol, triglycerides, and low-density lipoproteins by 8.3, 27.8, and 9.3%, respectively. The food additive is recommended for the planned course use in patients with MS.

[Use of biologically active supplement containing omega-3 polyunsaturated fatty acids in patients with type 2 diabetes mellitus]. / Primenenie biologicheski aktivnoi dobavki, soderzhashchei polinenasyshchennye zhirnye kisloty semeistva omega-3, u bol'nykh sakharnym diabetom 2 tipa.

35 patients with diabetes mellitus type 2 complicated with hyperlipidemia, coronary heart disease were treated during 8 months with poseidonol--polyunsaturated fatty acid family omega 3 (PUFA--omega 3). It was shown that hypercoagulation and hypofibrinolysis in these patients occurred. Treatment with poceidonol increased plasminogen activator activity and decreased coagulation and reduced atherogenic index.

[Long-term administration of tyquanol in the treatment of patients with ischemic heart disease in combination with hyperlipidemia]. / Dolgosrochnoe primenenie Tykvéinola v lechenii bol'nykh ishemicheskoi bolezn'iu serdtsa v sochetanii s giperlipidemiei.

33 patients with coronary heart disease and hyperlipidemia took tyquanol for 6 months. Tyquanol is a complex of biologically active substances represented by polyunsaturated fatty acids, antioxidants and polyvitamins produced of dietary oils of the sea and plant origin. As a result of the treatment with tyquanol cholesterol, beta-lipoproteins, triglycerides lowered by 16.1, 11.1, 26.3%, respectively. Alphipoproteins rose by 8.8%. A negative effect on hepatic, renal functions or electrolyte composition was not observed.

[Administration of the omega-3 polyunsaturated fatty acid drug eiferol decreases alcohol motivation in albino rats by elevating the level of antibodies to alcohol dehydrogenase]. / Vvedenie preparata polinenasyshchennykh zhirnykh kislot semeistva omega-3 (éiferola) snizhaet alkogol'nuiu motivatsiiu u belykh krys, povyshaia uroven' antitel k alkogol'degidrogenaze.

The study experimentally assessed the approach proposed by the authors to lower alcohol motivation, which involves enhancement of a specific immunity at the stage of alcoholization when acetaldehydemodified ethanol exchange enzymes [alcohol dehydrogenase (ADH)] and acetaldehyde dehydrogenase may be expected to occur. Omega-3 polyunsaturated fatty acid (PUFA) drugs enhance the formation of autoantibodies to modified ADH and decrease the activity of ADH in the stomach and liver. At the same time, PUFA drugs can, under certain conditions, produce an anti-alcoholic activity and a positive effect on the psychoemotional status of animals after the ethanol deprivation period.

[Effects of tycveinol on the lipid transport system and hemostasis and fibrinolysis systems in patients with ischemic heart disease]. / Vliianie tykvéinola na lipidtransportnuiu sistemu i sistemy gemostaza, fibrinoliza bol'nykh ishemicheskoi bolezn'iu serdtsa.

33 patients with ischemic heart diseases, hyperlipidemia and blood hypercoagulation took tycveinol which consists of polyunsaturated fatty acids in a dose of 2.0 g twice a day for 2 months. A positive result manifested by a 14% drop in cholesterol, 29.7% in triglycerides, 21% in beta-lipoproteins. Alpha-lipoproteins rose by 26%, atherogenic index reduced from 3.24 to 1.89. Hypercoagulatory tension relieved due to normalization of euglobulin lysis, higher activity of plasminogen tissue activator, slow blood clotting. Side effects on hepatic or renal function, mineral metabolism were not observed.

[Influence polyunsaturated fatty acids omega-3 on coagulation and fibrinolysis systems in patients with NIDDM]. / Vliianie polinenasyshchennykh zhirnykh kislot omega-3-klassa na sistemy svertyvaniia krovi i fibrinoliza bol'nykh sakharnym diabetom tipa 2]

35 patients with diabetes mellitus type 2 complicated with hyperlipidemia, coronary heart disease, were treated for 8 months with poseidonol which incorporates essential polyunsaturated fatty acids and fat-soluble vitamins. Poseidonol reduced atherogenic index from 4.2 to 3.15. The study of blood coagulation and fibrinolysis points to signs of hypocoagulation and normalization of fibrinolytic activity against previous hypercoagulation and fibrinolysis depression.

Effect of eiconol enriched with omega-3 polyunsaturated fatty acids on rat behavior and alcohol motivation.

We evaluated the effect of eiconol containing polyunsaturated fatty acids on the behavior and alcohol motivation in rats. Administration of eiconol for 10 days to alcoholized rats against the background of ethanol deprivation produced a sustained suppression of alcohol motivation and corrected deprivation-specific behavior.

[Poseidonol in the treatment of hyperlipidemia in patients with type 2 diabetes mellitus]. / Poseidonol v lechenii giperlipidemii u bol'nykh sakharnym diabetom tipa 2.

Thirty five patients with type 2 diabetes mellitus complicated by cardiovascular diseases received 6-month treatment for hyperlipidemia with poseidonol that represents a set of essential polyunsaturated fatty acids. This yielded good results as 11.6%-, 30%- and 24.2% decreases in the levels of cholesterol, triglycerides, and beta-lipoprotein, respectively, and a 9.9%- increase in alpha-lipoprotein. Lipid metabolism became normal in 34.2% of the patients, the remaining had positive changes as normalization of 1 to 3 lipid parameters. The tolerance of the agent was good, no negative effect on hepatic and renal functions was found.

Aminopeptidase PC from the hepatopancreas of the Kamchatka crab Paralithodes camtshatica.

Homogeneous aminopeptidase PC was isolated with yield 67% and purification degree 237 from the hepatopancreas of the Kamchatka crab Paralithodes camtshatica by ion-exchange chromatography on DEAE-Sepharose, hydrophobic chromatography on Phenyl-Sepharose, and gel-filtration on Sephadex G-150. The enzyme is a homodimer with a molecular mass 220 kD (110 x 2). Aminopeptidase PC has pI = 4.1. It hydrolyzes Leu-pNA optimally at pH 6.0 and at the optimum temperature 36-40 degrees C; in the presence of Ca2+ the enzyme is stable at pH 5.5-8.0. Aminopeptidase PC is activated by Ca2+, Mg2+, and Fe2+; it is completely inhibited by EDTA, o-phenanthroline, and bestatin. The enzyme contains four Zn atoms per molecule and is therefore a metalloaminopeptidase. The aminopeptidase PC can effectively cleave N-terminal Arg and Lys residues as well as Leu, Phe, and Met residues. Km and kcat values for hydrolysis of Leu-pNA were 0.075 mM and 0.19 sec-1 and for hydrolysis of Arg-pNA 0.078 mM and 0.48 sec-1, respectively. D-Amino acid residues cannot be cleaved. Thus, aminopeptidase PC of the Kamchatka crab has a mixed substrate specificity which is characteristic of some microbe aminopeptidases. Its N-terminal sequence ESVEIELPEGLSPLV is 46% coincident with that of yeast vacuolar aminopeptidase YSCA.

[Poseidonol in the treatment of patients with hyperlipidemia]. / Poseidonol v lechenii bol'nykh s giperlipidemiei.

An antilipidemic effect of poseidonol was tested in 123 patients with ischemic heart disease, stable angina of effort (II-III functional class), 40 patients with secondary pyelonephritis and 40 patients with chronic hepatosis-hepatitis associated with chronic alcoholism or diabetes mellitus. The patients had also hyperlipidemia. A positive effect of poseidonol was established: cholesterol content fell by 25.4, 20.6 and 18.7%, respectively; triglycerides level decreased by 63, 42 and 21.4%, respectively; B-lipoproteins fell by 55.5, 36.9 and 13.1%, respectively. High density lipoproteins rose by 40.2, 27 and 69.9%, respectively. No adverse effects either on the liver or kidney functions were observed.

[Effect of the proteolytic substance morikrase on the status of scar tissue under in vitro conditions]. / Deistvie proteoliticheskogo preparata morikrazy na sostoianie rubtsovoi tkani v usloviiakh in vitro.

Proteolytical preparation moricrasa was isolated from hepatopancreas of Paralithodes camtshatic. Moricrasa and its fractions were used for lysis of cicatricial tissue under in vitro conditions. It was found that moricrasa in concentrations 0.5 mg/ml and 2 mg/ml lysed the cicatrix during 72 and 14-16 hours, respectively. The fraction consisted of collagenolytic proteinase PC and trypsin revealed maximal activity of fraction of moricrasa was absent.