[The principles of analysis on multicomponent fluorescence spectra of proteins. Temperature and pH-induced transitions in tetrameric melittin].

Structural transitions in the tetrameric melittin from bee venom in 2 M KCl induced by variations of pH (from 0.7 to 12.0) and temperature (from 2 to 95 degrees C) have been studied. The pH and temperature ranges of structural changes and the zones of emission quenching of discerning tryptophan classes were revealed. The analysis of the temperature dependence of monotonic changes of spectral maximum positions and relative fluorescence yields allowed one to discriminate the zone of structural transitions in the tetramer from that of temperature quenching due to the thermal activation of fluorophore collisions with neighboring quenching groups in protein. Based on the new and earlier published results, some advantages and modes of using the method of component analysis of protein tryptophan spectra were summarized to determine the main characteristics of physicochemical transitions in proteins.

[Calcium-dependent interaction of transducin with calmodulin-sepharose].

It has been shown by affinity chromatography on calmodulin-sepharose that transducin, a G protein of bovine retinal rod outer segments interacts with Ca(2+)-calmodulin. This result assumes that the main part of calmodulin in dark retinal rod outer segments is associated with transducin. It has been suggested that photoactivation of retinal rods induces changes in intracellular calmodulin concentration, which may be one of the steps involved in the light adaptation of photoreceptor.

[The component analysis of tryptophan fluorescence spectra of melittin during its oligomerization].

The oligomerization of melittin with increasing ionic strength and protein concentration was investigated using the methods of decomposition of its tryptophan fluorescence spectra into "elementary" log-normal components. At high ionic strength (up to 2 M KCl), the emission spectra of tetrameric melittin are well described as the sum of two log-normal components, suggesting the presence of tryptophan residues in two sorts of environment with greatly differing polarity. Measurements of fluorescence spectra by iodide showed that these two spectral components possess different Stern-Volmer constants, that is, the tryptophans emitting them have different solvent accessibility, which does not correlate with the crystallographic structure of tetrameric melittin. Moreover, in the oligomerization transition induced by ionic strength, the tetrameric intermediate is formed, which has log-normal spectral components with relative contributions differing from those in 2 M KCl.

pH-induced transition and Zn2+-binding properties of bovine prolactin.

A pH-induced conformational transition was found in bovine prolactin within the physiologically significant pH region from 6.5 to 8.5. The thermal stability of prolactin at pH 6.5 is essentially higher than at pH 8.5. Bovine prolactin binds zinc ions with an apparent association constant of 2 x 10(5) M(-1) at pH 6.5 and 1 x 10(4) M(-1) at pH 8.5. The pH dependence of both thermal stability and zinc binding surrounding the pKa of histidine suggests that these residues plays a key role in the structural integrity of bovine prolactin.

Interactions of alpha-lactalbumins with lipid vesicles studied by tryptophan fluorescence.

Complexes of alpha-lactalbumin (alpha-LA)1 with dimyristoylphosphatidylcholine (DMPC) or dipalmitoylphosphatidylcholine (DPPC) liposomes at pH 8 and at pH 2 have been obtained by means of gel filtration. Thermal denaturation of alpha-LA complexes of DMPC or DPPC at pH 8 was found to depend on the saturation of protein by metal cations. The intrinsic fluorescence of DMPC-alpha-LA and DPPC-alpha-LA was sensitive to two thermal transitions. The first transition corresponded to the Tc of the lipid vesicles, while the second transition arose from the denaturation of the protein. Fluorescence spectrum position suggested that at low temperature tryptophan accessibility increases upon protein-DMPC or protein-DPPC association. At temperatures above the protein transition (70 degrees C) tryptophan appears to interact significantly with the apolar phase of DMPC and DPPC, evidenced by spectral blue shifts. Whereas the free protein at pH 2 adopts the molten globule (MG) state and is characterized by the absence of a thermal transition, the rapidly-isolated DMPC-alpha-LA complex was characterized by the appearance of a distinct fluorescence thermal transition between 50 and 60 degrees C. This result is consistent with a model of a partially-inserted form of alpha-LA which may possess some degree of tertiary structure and therefore unfolds cooperatively.

Blockers of Ca2+ channels in the plasmalemma of perfused Characeae cells.

Ionic currents in the plasmalemma of perfused Nitella syncarpa cells identified as currents through Ca2+ channels were registered for the first time. The effect of 1,4-dihydropyridine derivatives (nifedipine, nitrendipine, riodipine) and phenylalkylamines (verapamil, D600) as well as the agonist CGP-28392 on the Ca2+ channels in the plasmalemma of perfused cells of Nitellopsis obtusa and Nitella syncarpa have been studied. A blocking effect of 1,4-dihydropyridine derivatives and phenylalkylamines on the plasmalemma Ca2+ channels has been detected. Phenylalkylamines have been found to block both inward and outward Ca2+ currents. The activating effect of the agonist CGP-28392 on the Ca2+ channels of plasmalemma has been shown.

Calcium-regulated interactions of human alpha-lactalbumin with bee venom melittin.

Affinity chromatography, fluorescence and circular dichroism spectroscopy methods have been used to study the interaction of melittin, a 26-residue peptide from bee venom, with Ca2(+)-binding alpha-lactalbumin from human milk. It has been revealed that melittin binds to the apo- and acidic states of alpha-lactalbumin while the presence of Ca2+ makes the interaction essentially weaker. The association constant for the complex of melittin with apo-alpha-lactalbumin determined from spectropolarimetric melittin-titration data is 2 X 10(7) M-1. The complexation of alpha-lactalbumin with melittin decreases its affinity to Ca2+ by three orders of magnitude. The interaction of apo-alpha-lactalbumin with melittin causes some changes in the environment of its aromatic amino acid residues and drastically alters the conformation of melittin, increasing its alpha-helical content but leaving its single tryptophan residue accessible to water. In the case of the acidic state of alpha-lactalbumin the interaction does not induce an increase in alpha-helical content of melittin.

[Endoscopic hemostasis in pulmonary hemorrhage]. / Endoskopicheskii gemostaz pri legochnom krovotechenii.

As the result of applying endoscopic occlusion of the bronchus in complex treatment of 95 patients with pulmonary, predominantly massive, hemorrhage recovery was achieved in 77.9% of cases. Twenty-one patients died. Hemorrhage was the direct cause of death of 5 patients, 16 patients died from advancement of the principal disease, background conditions, and complications of the postoperative period not associated with pulmonary bleeding.

[Extracellular RNAase Pch2 of Penicillium chrysogenum 152A: purification, specificity and physico-chemical properties]. / Vnekletochnaia RNKaza Pch2 Penicillium chrysogenum 152A. Ochistka, spetsifichnost' i fiziko-khimicheskie svoistva.

Acid RNAase Pch2 was isolated from a filtrate of the cultural fluid of the fungus Penicillium chrysogenum 152A and purified to homogeneity. An analysis of RNAase Pch2 action on RNA and synthetic substrates showed that the enzyme can be attributed to non-specific true ribonucleases (ribonucleate-3'-oligo-nucleotide hydrolase, EC 3.1.4.23). The maximal effect of the enzyme on RNA is observe at pH 4.5 and 55 degree. The RNAase Pch2 is not activated by bivalent metal ions, p-chloromercurybenzoate or beta-mercaptoethanol and is reversibly inactivated by 8 M urea. The enzyme molecule consists of 332 amino acid residues; its molecular weight is 36160, the isoelectric point lies at 5.2.

[Amino acid composition and proton capacity of pea chloroplast thylakoid membranes]. / Aminokislotnyi sostav i protonnaia emkost' tilakoidnykh membran khloroplastov gorokha.

The amino acid composition and pH dependence of proton capacity of pea chloroplast thylakoid membranes have been studied. The thylakoid membranes possess a high content of amino acids, whose functional groups can either bind or split off protons depending on pH of the reaction mixture. The number of amino acids with proton-accepting functional groups per mg of chlorophyll significantly exceeds the proton capacity of the native membranes. It is concluded that the buffer capacity of thylakoid membranes is predetermined by the proton-accepting groups of protein components which are largely blocked within the native structure of the membranes. The protein/chlorophyll weight ratio for pea chloroplast thylakoid membranes is 3.22 +/- 0.05.

[Thermodynamic study of guanylspecific ribonuclease C2]. / Termodinamicheskoe issledovanie guanilspetsifichnoi ribonukleazy C2.

Guanylspecific RNAse C2 has been studied thermodynamically using a microcalorimetry technique. It is shown that at ph varying from 5,0 to 8,0 the RNAse C2 solution in a cacodylate buffer contains about 30% of monomeric and about 70% of dimeric forms of protein, each of them melting as a single cooperative system. Moreover, studies of the influence of phosphate ions have shown that they stabilize the system by 3-4 degrees and transform it into a monomeric form.

[Amino acid composition and chemical functional group modification of protease (TI-Ajl) inhibitor from Actinomyces janthinus 118]. / Aminokislotnyi sostav i khimicheskaia modifikatsiia funktsional'nykh grupp ingibitora proteaz (IT-Ajl) iz Actinomyces janthinus 118.

The amino acid composition of the protease (TI-Ajl) inhibitor from Actinomyces janthinus 118 has been determined. It was shown that the TI-Ajl, S-SI and plasminostreptin inhibitors of trypsin and subtilisin have a number of common features: number of double bonds, tryptophane and tyrosine residues, prevalence of the acidic amino acids over the basic ones, ets. The effect of chemical modification of amino groups, arginine, tyrosine and methionine residues on the inhibitory activity of TI-Ajl was studied. The data obtained are indicative of the presence of two active centers of the inhibitor. The antitrypsin center contains a lysine residue.

[Photo-oxidation and carboxymethylation of guanylribonuclease Pch1]. / Fotookislenie i karboksimetilirovanie guanilspetsifichnoi ribonukleazy Pch1.

The effect of photo-oxidation and carboxymethylation on the activity of RNAse Pch1 has been studied. Photoinactivation in the presence of rose bengal results in a selective oxidation of two histidine residues. The process is inhibited by the nucleotide substrate analogs. This suggests that one or two imidazole groups may be localized in the active site of RNAse Pch1. The pH dependence of the enzyme inactivation by bromoacetic acid is indicative of the contribution of a functional group with pKa 4,0, presumably of a beta- or gamma-carboxyl group of dicarbonic amino acid. The reaction is inhibited by the substrate analogs 2'(3')-GMP and 2'(3')-AMP. The data on the similarity of active sites in several guanyloribonucleases are discussed.

Influence of the conditions of growth on the chemical composition and activity of the enzymes of the cell envelope of the yeast Candida tropicalis IBFM-303.

The article is devoted to a study of the changes in the activity and chemical composition of the enzymes of the cell envelope of the yeast Candida tropicalis IBFM-303 during growth on n-alkanes and glucose, as well as the transport of n-alkanes through the cell membrane and the redistribution of the cell contents in the process of budding on the indicated carbon sources.