ABSTRACT
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.
Subject(s)
Androstanes/metabolism , Ascomycota/metabolism , Cortodoxone/metabolism , Pregnanes/metabolism , Ascomycota/genetics , Ascomycota/ultrastructure , Biotransformation , Hydroxylation , MutationABSTRACT
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.
Subject(s)
Adjuvants, Immunologic/metabolism , Dehydroepiandrosterone/metabolism , Fungi/metabolism , Adjuvants, Immunologic/pharmacology , Catalysis , Dehydroepiandrosterone/pharmacology , Fungi/growth & development , HydroxylationABSTRACT
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).
Subject(s)
Arthrobacter/metabolism , Hydroxysteroid Dehydrogenases/metabolism , Nocardiaceae/metabolism , Steroids/metabolism , Arthrobacter/classification , Cortodoxone/analogs & derivatives , Cortodoxone/chemistry , Cortodoxone/metabolism , Nocardiaceae/classification , Phylogeny , RNA, Bacterial/analysis , RNA, Ribosomal, 16S/analysis , Steroids/chemistryABSTRACT
A screening for carbon sources revealed ethanol to be the best substrate for condensed Nocardioides sp. culture. A strategy achieving the maximum (21 g/l) yield of biomass was developed for the control over the condensed fed-batch culture production. This control based on the algorithm ExpoDense should be predetermined in the first phase and adaptive in the second phase of two-phase process of condensed culture production.
