[Bioconversion of C19- and C21-steroids with parent and mutant strains of Curvularia lunata].

Regio- and stereospecificity of microbial hydroxylation was studied at the transformation of 3-keto-4-ene steroids of androstane and pregnane series by the filamentous fungus of Curvularia lunata VKMF-644. The products of the transformations were isolated by column chromatography and identified using HPLC, mass-spectrometry (MS) and proton nuclear magnetic resonance (1H NMR) analyses. Androst-4-ene-3,17-dione (AD) and its 1(2)-dehydro- and 9alpha-hydroxylated (9-OH-AD) derivatives were hydroxylated by the fungus mainly in position 14alpha, while 6alpha-, 6beta- and 7alpha-hydroxylated products were revealed in minor amounts. At the transformation of C21-steroids (cortexolone and its acetylated derivatives) the presence of 17-acetyl group was shown to facilitate further selectivity of 11beta-hydroxylation. Original procedures for protoplasts obtaining, mutagenesis and mutant strain selection have been developed. A stable mutant (M4) of C. lunata with high 11beta-hydroxylase activity towards 21-acetate and 17alpha,21-diacetate of cortexolone was obtained. Yield of 11beta-hydroxylated products reached about 90% at the transformation of 17alpha, 21-diacetate of cortexolone using mutant strain M4.

[Dihydroxylation of dehydroepiandrosterone in positions 7alpha and 15alpha by mycelial fungi].

The ability of 485 fungal strains is studied for catalysis of the process of 7alpha, 15alpha-dihydroxylation of dehydroepiandrosterone (DHEA, 3alpha-hydroxy-5-androstene-17-one), a key intermediate of the synthesis of physiologically active compounds. The ability for the formation of 3alpha, 7alpha, 15alpha-trihydoxy-5-androstene-17-one (7alpha, 15alpha-di-OH-DHEA) was found for the first time for representatives of 12 genera, eight families, and six orders of ascomycetes, eight genera, four families, and one order of zygomycetes, one genus, one family, and one order of basidiomycetes, and four genera of mitosporous fungi. The most active strains are found among genera Acremonium, Gibberella, Fusarium, and Nigrospora. In the process of transformation of DHEA (2 g/l) by strains of Fusarium oxysporum RKM F-1600 and FGibberella zeae BKM F-2600, the molar yield was 63 and 68%, respectively. Application of the revealed active strains of microorganisms opens prospects for the efficient production of key intermediates of synthesis of modern medical preparations.

[The 1(2)-dehydrogenation of steroid substrates by Nocardioides simplex VKM Ac-2033D]. / 1(2)-degidrirovanie steroidnykh substratov shtammom Nocardioides simplex BKM Ac-2033D.

The bacterium formerly known as Arthrobacter globiformis 193 has high 1(2)-dehydrogenase activity toward pharmaceutically important steroids, 9(11)-dehydrocortexolone in particular. The complex analysis of the morphostructural, physiological, biochemical, and phylogenetic properties of this bacterium allowed us to reclassify it into Nocardioides simplex (N. simplex VKM Ac-2033D).

[Effect of emulsifiers released by yeasts multiplying on n-alkane media on their growth and microscopic organization]. / Vliianie émul'gatorov, vydeliaemykh drozhzhami, razmnozhaiushchimisia na sredakh s n-alkanami, na ikh rost i mikroskopicheskuiu organizatsiiu.

Bioemulsifiers and ether extracts from the cultural broth of yeast cells were used to study their effect of Candida lipolytica growth and ultrastructural organisation. When the emulsifiers and ether extracts were added to the growth medium, the lag phase was reduced but the growth rate did not change. The ether extracts increased the growth rate of C. lipolytica 374/2 and the final biomass yield of C. lipolytica 704. The ultrastructural organisation of C. lipolytica 374/2 cells changed under the action of the bioemulsifier added to the growth medium.

[Alcohol oxidation by Candida guilliermondii yeasts grown on hexadecanol]. / Okislenie spirtov drozhzhami Candida cuilliermondii, vyrashchennymi na geksadekanole.

A number of enzyme systems involved in the first steps of hexadecane oxidation can be induced by hexadecanol, an intermediate product of hexadecane degradation. It has also been found that, in Candida guilliermondii cells and in their mitochondrial fraction, an oxidase system is induced when the cells are grown on hexadecanol. This system is similar to that in cells grown on hexadecane; it oxidises higher alcohols at a high rate and is not inhibited by the inhibitors of the man phosphorylating respiration chain. The membrane-bound alcohol dehydrogenase and aldehyde dehydrogenase activities resistant to pyrazole, an inhibitor of cytosol ethanol dehydrogenase, are induced together with the oxidase activity when the cells are grown on hexadecanol as well as on hexadecane. The oxidation of higher alcohols by whole cells is entirely inhibited by azide although their oxidation by mitochondria is resistant to the action of azide; apparently, azide inhibits the transport of alcohols into the cell.

[Effect on the orientation of the membrane vesicles of Escherichia coli of various methods of cell disintegration]. / Vliianie na orientatsiiu membrannykh vezikul u Escherichia coli razlichnykh sposobov dezintegratsii kletok.

As was demonstrated using the Con-A polymer, membranous fractions prepared by various cell disintegration procedures are a heterogeneous population. The population includes right side out vesicles and inside out vesicles whose proportion depends on the procedure of disintegration. The orientation of these vesicles was studied by electron microscopy, their ATPase activity was assayed by cytochemical techniques, and the morphology of the vesicles was also investigated. The authors discuss the possible effect of Con-A on the reorganisation of membranes and the activity of ATPase.

[Yield of Chlamydia from an infected cell studied by scanning electron microscopy]. / Izuchenie vykhoda khlamidii iz infitsirovannoi kletki metodom skaniruiushchei élektronnoi mikroskopii.

The study of the monolayer cell culture L-929, infected with strain CP-1 isolated from a patient with Reiter's syndrome, by means of scanning electron microscopy demonstrated that the release of chlamydiae from infected cells occurred in two ways: (1) by active ejection, probably as the consequence of the local rupture of the host cell, and (2) by more quiet "eruption" due to rupture in the plasmolemma . Elementary chlamydial bodies released from the cell had a spherical form, and reticular bodies had an irregularly spherical form, often with recesses and digitiform protrusions.

[Dependence of the efficiency of the fractionation of summary E. coli cell membranes on the method of their disintegration]. / Zavisimost' éffektivnosti fraktsionirovaniia summarnykh membran kletok E. coli ot sposoba ikh dezintegratsii.

The efficiency of fractionation in the sucrose density gradient of E. coli cell membranes obtained after cell disintegration by ultrasonic, ballistic and extrusion methods was measured. The purity of individual membrane fractions was estimated by electron microscopy and polyacrylamide gel electrophoresis. The application of ballistic disintegration and solid state extrusion did not separate inner and outer membranes. Cell disintegration by means of ultrasonic treatment and liquid state extrusion allowed reproducible separation of membranes of the two types with a sufficiently high degree of purity.

[Structural and biochemical characteristics of Escherichia coli cells with various levels of secretable alkaline phosphatase]. / Strukturno-biokhimicheskie osobennosti kletok Escherichia coli s raznym urovnem sinteza sekretiruemoi shchelochnoi fosfatazy.

The biochemical and structural peculiarities of Escherichia coli cells with different rates of synthesis of secreted alkaline phosphatase were studied under the conditions of repression, derepression and constitutive synthesis. The enzyme synthesis was shown to correlate with the redistribution of free and membrane-bound ribosomes in the cells, with an increase in their proportion, and with a reconstruction of the cell ultrastructural organization.

[Cytology of the dynamics of capsule formation in yeasts]. / Tsitologiia dinamiki obrazovaniia kapsul u drozhzhei.

Changes in the dimensions and structure of capsules were examined using optical and electron microscopy in the yeast Cryptococcus magnus producing extracellular polysaccharides in the course of its batch cultivation. The mean thickness of capsules was minimal in the period of active budding, but is rapidly increased later on. The dimensions of capsules, as well as the distribution density of capsule fibrils, were maximal at the beginning of the stationary growth phase. The results indicate that the intensive process of capsule formation occurs in formed buds, young separated cells. They contain the greatest number of vesicles with fibrillar material moving from the centre of the cell toward its periphery. The vesicles are absent in the stationary growth phase when the capsules thicken on more. In the phase of dying off, the capsule size and the density of capsule fibres decrease because the capsule material dissolves in the medium. These findings are discussed in relation to capsule biosynthesis and production of extracellular polysaccharides by yeasts.

[Submicroscopic surface organization of Candida lipolytica cells growing on hexadecane]. / Submikroskopicheskaia organizatsiia poverkhnosti kletok Candida lipolytica pri roste na geksadekane.

The ultrastructure of the cell wall surface was studied in two Candida lipolytica strains, IBPhM-160 (704) and 1, during their growth on hexadecane, using the technique of ultrathin sections. Linear three-layer membranes, membrane vesicles and complicated membrane complexes were shown to appear on the surface of the cell walls when the yeast strains were grown on hexadecane. The rate of membrane formation was greater in the cells of strain 1, and increased in the both strains by the stationary growth phase. Since no cells underwent lysis at any of the growth phases studied, the formation of membrane complexes on the yeast cell wall should reflect the reconstruction of the surface of morphologically intact cells grown on hexadecane.

[Cytobiochemical characteristics of Candida boidinii in continuous culture on methanol and HCO3-]. / Tsitobiokhimicheskie osobennosti Candida boidinii pri nepreryvnom kul'tivirovanii na metanole i HCO3-.

The cytological and biochemical characteristics of the yeast Candida boidinii KDI were studied under the conditions of continuous cultivation in media containing methanol in the presence of exogenous bicarbonate and without it. Bicarbonate addition was found to cause a decrease in the activity of catalase, a reorganization of the peroxisomes, and an increase in the activity of pyruvate and phosphoenolpyruvate carboxylases, the enzymes involved in heterotrophic assimilation of carbon dioxide.

[Electron microscopic study of Bacillus polymyxa in the developmental process and antibiotic formation]. / Elektronno-mikroskopicheskoe izuchenie Bacillus polymyxa v protsesse razvitiia i obrazovaniia antibiotika.

The purpose of this work was to study growth of Bacillus polymyxa 153, biosynthesis of polymyxin by it, the formation of spores, and the fine structure of the cells. The antibiotic content was low in the cells and in the cultural broth during the first hours of growth and sharply increased later under the conditions of the medium ensuring the "vegetative" type of cultural growth. The cell wall reorganized, a periplasmic space appeared, and the membranous and mesosomal apparatus became more complicated in the cells actively synthesizing the antibiotic. The antibiotic content was higher in the cells belonging to the "spore" type of growth at the beginning of mass spore formation. Further cell development involved endospore differentiation; a complicated membranous apparatus did not appear, and biosynthesis of the antibiotic stopped.

[Yeast capsule ultrastructure]. / Ul'trastruktura kapsul drozhzhei.

The ultrastructure of capsules was studied in 12 yeast species producing extracellular polysaccharides and belonging to ascomycetous, basidiomycetous and asporogenous organisms using the method of ultrathin sections. On the whole, the structure of capsules was similar in these organisms: the capsules are formed by fibrils radially coming out of the outer surface of the cell wall. Unlike those of basidiomycetous yeasts, the capsules of ascomycetous organisms are characterized by a more ordered orientation of fibrils which run strictly parallel to one another. In the capsules of basidiomycetous organisms, fibrils usually interweave, often producing a fibrillar network; in certain species, fibrils stick together forming thick threads on the periphery of the capsule. Moreover, as a rule, these yeasts possess a distinct bright zone (halo) above the outer surface of the cell wall. These results as well as data available about the chemical composition and functions of the capsule indicate that there are radical differences the latter and the cell wall which make it possible to regard the capsule as an individual organelle of the yeast cell rather than as part of the cell wall.

[Morphologic characteristics of the large bodies in cultures of the L-forms of bacteria according to scanning electron microscope studies]. / Morfologicheskaia kharakteristika bol'shikh tel v kul'turakh L-form bakterii po dannym skaniruiushchei élektronnoi mikroskopii.

Large bodies appear at the time of protoplast and spheroplast formation and are revealed at all the L-transformation stages and at the initial stage of reversion. They can be represented both by a single giant cell and by a conglomerate of different cells connected with one another. They are not only spheroid, but can be of the most varied shape, and structurally they are connected with other L-colony elements: filamentous structures, spheroid cells, elementary bodies and the so-called acellular material. At the early L-transformation stage the large bodies probably appear as a result of coalescence of lysed cells and represent polygenome formations. Elementary bodies and spheroid cells form within the large bodies and on their surface at the late stage of L-transformation. In case of reversion bacterial cells form from them.

[Morphologic characteristics of spheroid elements in a culture of bacterial L-forms according to scanning electron microscopy findings]. / Morfologicheskaia kharakteristika sharovidnykh élementov v kul'ture L-form bakterii po dannym skaniruiushchei élektronnoi mikroskopii.

The authors studied stable L-cultures of Proteus valgaris, Bac. subtillis, Staphylococcus aureus, Streptococcus pyogenes of group A, and also unstable cultures of the L-forms of Proteus vulgaris and Proteus vulgaris culture at the stage of spheroplasts. Spheroid cells proved to appear at the stage of spheroplasts, prevailed at the log phase in stable and unstable L-cultures, but were less frequent at the stationary phase. Cross section of L-colonies showed that they were located at the surface. The size of spheroid elements was from 1 to 5 micron; their surface was smooth or slightly wrinkled with numerous protrusions and individual sockets. The spheroid cells were distributed in the colonies freely, in clusters, or were connected to one another by anastomosis. Several methods of reproduction of spheroid cells are described, including equal and unequal binary fission, budding, and formation of elementary bodies within the cell. Morphological connection of spheroid cells with large bodies, filamentous structures and structureless matrix of the L-colony apparently pointed to their origin from the corresponding elements of the L-cultures.

[Study of streptococcal L forms in the scanning electron microscope. II. Dynamics of the development of structural elements of L colonies at different stages of growth]. / Izuchenie L-form streptokokka v skaniruiushchem élektronnom mikroskope. Soobshchenie II. Dinamika razvitiia strukturnykh élementov L-kolonii na raznykh stadiiakh rosta

The method of scanning electron microscopy showed that the L-colonies of streptococcus were formed by the spherical structures 0.1--1.5 micronm in diameter, elements of polygonal shape (large bodies) 10--30 micronm in size, filamentous structures 01--7 micronm in diameter and structureless matrix. A regular replacement of one form by another was observed in the process of the L-colonies development. Thus, the spherical elements appeared in the lag-phase, and polygonal elements were found mostly at the initial stages of the L-colonies formation; as to the filamentous structures -- they were present at all the developmental stages, but their diameter increased, and their structure and number changed at different growth phases. The spherical elements of the L-colonies formed evenly both on the structureless depth matrix of the colonies, on the filamentous structures in the form of buds on the "large bodies", and the disintegration of the latter. The role of the filamentous structures in the development of the L-colonies is discussed.