Molecular determinants of voltage-dependent slow inactivation of the Ca2+ channel.

Ba(2+) current through the L-type Ca(2+) channel inactivates essentially by voltage-dependent mechanisms with fast and slow kinetics. Here we found that slow inactivation is mediated by an annular determinant composed of hydrophobic amino acids located near the cytoplasmic ends of transmembrane segments S6 of each repeat of the alpha(1C) subunit. We have determined the molecular requirements that completely obstruct slow inactivation. Critical interventions include simultaneous substitution of A752T in IIS6, V1165T in IIIS6, and I1475T in IVS6, each preventing in additive manner a considerable fraction of Ba(2+) current from inactivation. In addition, it requires the S405I mutation in segment IS6. The fractional inhibition of slow inactivation in tested mutants caused an acceleration of fast inactivation, suggesting that fast and slow inactivation mechanisms are linked. The channel lacking slow inactivation showed approximately 45% of the sustained Ba(2+) or Ca(2+) current with no indication of decay. The remaining fraction of the current was inactivated with a single-exponential decay (pi(f) approximately 10 ms), completely recovered from inactivation within 100 ms and did not exhibit Ca(2+)-dependent inactivation properties. No voltage-dependent characteristics were significantly changed, consistent with the C-type inactivation model suggesting constriction of the pore as the main mechanism possibly targeted by Ca(2+) sensors of inactivation.     

Molecular tools for the study of calcium channels: synthesis and evaluation of biological activity of new derivatives of 4-aryl-1,4-dihydropyridine groups

The synthesis and biological activity of some novel analogs of the calcium channel blocker nifedipine, i.e., derivatives of 2.6-dimethyl-3.5-diethoxycarbonyl-4-(3-nitrophenyl)-1.4-dihydropyridine (DHP) were studied. One radioactive and two photoactivable DHP derivatives were obtained. DHP hemisuccinate was used to prepare an affinity matrix, DHP-Sepharose as well as a DHP-albumin conjugate; the latter was used for anti-DHP antibodies generation in rabbits. All novel DHP derivatives were obtained from a single key 3-hydroxycarbonyl DHP derivative, and they comprise a series of necessary tools for the study and isolation of membrane calcium channels.     

Organ blood flow and vessels of microcirculatory bed in fish

Information on density of fish capillary network and its permeability, peculiarities of geometry, morphology, and ultrastructure of vessels of microcirculation bed—arterioles, venules, capillaries—is presented. A great attention is paid to vasomotor reactions and their participation in redistribution of blood. Nervous and humoral mechanisms of control of tone of the vessel smooth muscle wall and voluminous blood flow are considered. Effects of environmental factors on processes of microcirculation in fish are discussed.     

Purification and characterization of dihydropyridine receptor from rabbit skeletal muscle

Digitonin and 3-[(3-cholamidopropyl)dimethylammonio]-2-hydroxy-1-propane sulfonate (Chapso) were used to solubilize the receptor of dihydropyridine calcium antagonists from the transverse tubule membranes of rabbit skeletal muscle. The receptor retained the ability for selective adsorption from either detergent extract by dihydropyridine-Sepharose. Incubation of the affinity resin with nitrendipine resulted in the elution of the receptor protein composed of two main polypeptides with molecular masses of 160 kDa and 53 kDa, as shown by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. Only these two subunits were found in the receptor preparation purified to a specific dihydropyridine-binding activity of 2500-2800 pmol/mg protein (60-70% purity) from digitonin-solubilized membranes by a combination of wheat-germ-agglutinin--Sepharose, anion-exchange and dihydropyridine-Sepharose chromatography steps. The individual subunits were isolated in dodecyl-sulfate-denatured form from the preparation of the receptor, enriched by a two-step large-scale procedure applied to Chapso-solubilized membranes. The 160-kDa subunit slowly changed its apparent molecular mass to 125 kDa upon disulfide bond reduction without formation of novel peptides. This finding implies that 160-kDa subunit is cross-linked by intramolecular S-S bridge(s). Chemical deglycosylation with trifluoromethanesulfonic acid showed that the carbohydrate content of large and small subunits accounted for 7.5% and 6.6% by mass, respectively. The dihydropyridine receptor subunits are glycosylated through N-glycoside bonds only. In their ratio of polar to hydrophobic amino acid residues in the amino acid composition of the receptor subunits, these polypeptides behave rather as peripheral proteins. It is suggested that the main portion of polypeptide chains is located outside the membrane in contact with solvent.     

A new role for PGRP-S (Tag7) in immune defense: lymphocyte migration is induced by a chemoattractant complex of Tag7 with Mts1

PGRP-S (Tag7) is an innate immunity protein involved in the antimicrobial defense systems, both in insects and in mammals. We have previously shown that Tag7 specifically interacts with several proteins, including Hsp70 and the calcium binding protein S100A4 (Mts1), providing a number of novel cellular functions. Here we show that Tag7–Mts1 complex causes chemotactic migration of lymphocytes, with NK cells being a preferred target. Cells of either innate immunity (neutrophils and monocytes) or acquired immunity (CD4⁺ and CD8⁺ lymphocytes) can produce this complex, which confirms the close connection between components of the 2 branches of immune response.     

Mutability of the LYS2 gene in diploid saccharomycete yeasts. I. The occurrence of spontaneous mutants and mutants induced by x-ray and ultraviolet radiation

More than 3000 spontaneous and induced lys2 mutants were obtained in haploid and diploid strains of yeast Saccharomyces. The ability to utilize alpha-aminoadipate was used for lys2 mutant screening. The spontaneous and induced mutation rates were measured in haploid and diploid strains. Mitotic segregation of pho1 marker linked to LYS2 was studied in lys2 mutants obtained in diploid strains. Fertility of diploid lys2 mutants was tested. The conclusion to be drawn from the data presented is that mutations appeared in one of two homologous chromosomes and then segregated by mitotic homozygotization.     

Prognosis of the incidence of hemorrhagic fever with the renal syndrome in the Primorsk region]

A study was made of a possibility of prognostication of the number of patients (morbidity per 100 000 residents) with hemorrhagic fever with the renal syndrome in conditions of the Primorsk region. The main factors characterizing this disease were analyzed with the use of the "Minsk-22" computer; these were dynamics of the populations of murine rodents, metereological data, morbidity. A method of prognostication of the number of patients (morbidity) with hemorrhagic fever and a renal syndrome in the Primorsk region for the current year was elaborated.     

Tissue Peculiarities of Energy Metabolism Enzyme Activity and ATP Content In Black Sea Ruff Scorpaena porcus (Scorpaenidae)

Journal of Ichthyology - The activities of cytosolic oxidoreductases (malate and lactate dehydrogenases) and the level of ATP production in the hypoxic resistive tissues of Scorpaena porcus...     

Methemoglobin and the Activities of Catalase and Superoxide Dismutase in Nucleated Erythrocytes of Scorpaena porcus (Linnaeus, 1758) under Experimental Hypoxia (in vitro)

The effect of hypoxia on nucleated red blood cells of the black scorpionfish (Scorpaena porcus) was studied in vitro. Deep hypoxia (the oxygen concentration was less than 1 mg O₂ L^–1; the norm was 7–8 mg O₂ L^–1) led to the transition of a part of the hemoglobin molecules to the ferric state (methemoglobin). The maximum increase in the concentration of methemoglobin was 32%. The accumulation of methemoglobin in red blood cells was accompanied by an increase in the activity of catalase and superoxide dismutase and a decrease in the content of reactive oxygen species in the cytoplasm of cells. It was shown that the formation of methemoglobin did not cause damage to the cytoplasmic membranes of red blood cells. The percentage of red blood cell lysis in deoxygenated (less than 1.0 mg O₂ L^–1) suspensions quantitatively coincided with the control values.

     

Qualitative composition of carotenoids, catalase and superoxide dismutase activities in tissues of the bivalve mollusc Anadara inaequivalvis (Bruguiere, 1789)

Using high performance liquid chromatography, UV-VIS spectra and mass-spectra (FAB MS), 7 carotenoid species were identified in tissues of the bivalve mollusc Anadara inaequivalvis (Bruguiere, 1789): trans- and cis-pectenolon, alloxanthin, pectenol A, β-carotene, zeaxanthin, and diatoxanthin. Their quantitative ratio in hepatopancreas, gills and foot of animals were determined. A negative correlation (R ² is about 0.9) was revealed between tissues content of a series of carotenoids (trans- and cis-pectenolon, zeaxanthin, alloxanthin, and diatoxanthin) and activity of antioxidant enzymes (catalase and superoxide dismutase). The existence of competitive interrelations between these molecular systems is proposed and underlying causes are discussed.