Igor Belič

Hydroxylation of steroids with 11α-hydroxylase of Rhizopus nigricans

Abstract Three groups of 3-keto-4-ene steroids with different side chains were used as substrates for the induced 11α -hydroxylase of Rhizopus nigricans . The highest total bioconversion as well as the highest yield of 11α-hydroxylated product is found using progesterone as substrate. By changing the polarity of the side chain, much higher yields of 6β- and 7β-hydroxylated products relative to 11α-hydroxylated product are obtained. Our results thus provide evidence for the importance of the side chain in steroid-enzyme interactions.

Steroid hormone signalling system and fungi

Three components of the steroid hormone signalling system, 17 beta-hydroxysteroid dehydrogenase, androgen binding proteins and steroid hormone signalling molecule testosterone were determined in the filamentous fungus Cochliobolus lunatus for the first time in a fungus. Their possible role in C. lunatus is discussed in comparison with their role in mammalian steroid hormone signalling system. The results are in accordance with the hypothesis, that the elements of primordial signal transduction system should exist in present day eukaryotic microorganisms.

11β-Hydroxylation of steroids by Cochliobolus lunatus

Abstract The substrate specificity of 11β-hydroxylase of Cochliobolus lunatus was studied and a close parallelism to the results obtained with 11α-hydroxylase of Rhizopus nigricans was observed. It was found that the cell wall does not differentiate between the steroid substrates used and the absence of the cell wall increases the bioconversion.

Microbial dehydrogenation of tomatidine

Abstract Tomatidine (tomatanin-3β-ol) was dehydrogenated by Nocardia restrictus to tamatanin-3-one, 1-tomatenin-3-one, 4-tomatenin-3-one and 1,4-tomatidien-3-one. No degradation of 1,4-tomatadien-3-one by Nocardia restrictus could be achieved.

Hydroxylation of steroids with nonpolar side chains with 11α-hydroxylase of Rhizopus nigricans

Abstract Steroids with nonpolar side chains with 2, 4 and 8 C atoms were used as substrates for the 11α-hydroxylase of Rhizopus nigricans. Their bioconversion was compared to that of progesterone, which was found to be far the best substrate giving the highest total bioconversion. 3-keto-4-ene steroids with nonpolar side chains were converted to their hydroxylated products in a small yield or not at all. The absence of an oxygen function in the side chain did not affect the regio-specificity of the hydroxylation, but resulted in a much lower total bioconversion. The strong effect of the oxygen function and of the length of the side chain on hydroxylation with the 11α-hydroxylase of Rhizopus nigricans was demonstrated.

Epimerisation of 3β-ol to 3α-ol steroid alkaloids by Nocardia restrictus

Abstract (22S,25S)-5α-tomatanin-3β-ol, N-acetyl-(22S,25S)-5α-tomatanin-3β-ol, (22R,25R)-5α-tomatanin-3β-ol and (22R,25S)-22,26-epimino-5α-cholestane-3β,16β-diol are transformed by Nocardia restrictus into corresponding 3α-ol compounds with yields from 70 to 5%.

Microbial dehydrogenation of steroid alkaloids with tertiary amino groups

Abstract N-methyl-5α-tomatanin-3β-ol, 5α-conanin-3β-ol and demissidine are dehydrogenated by Nocardia restrictus to the corresponding 1,4-diene-3-keto derivatives. N-acetyl-5α-tomatanin-3β-ol is dehydrogenated mainly to its 3-keto derivative, the 1,4-diene-3-keto derivative being obtained in only 1% yield.

Bioconversion of steroid glycosides by Nocardia restricta

The bioconversion of steroid alkaloid tomatine by Nocardia restricta yields the conjugate with lactic acid. We studied the bioconversion of some steroid glycosides without a nitrogen atom in the molecule to determine the effect of the nitrogen atom. The glycosides were of three different types: sterol glycosides, bufadienolide rhamnoside and steroid saponine. The results of bioconversions showed that Nocardia restricta converts steroid glycosides differently according to the sugar bound to the steroid aglycone. It can be concluded that in the absence of a nitrogen atom in the steroid molecule no conjugation with lactic acid by Nocardia restricta occurs.

Microbial dehydrogenation of dihydrotomatidines

Abstract (22S, 25S)-22, 26-Epimino-5α-cholestane-3β, 16β-diol and (22R, 25S)-22, 26-epimino-5α-cholestane-3β, 16β-diol were dehydrogenated by Nocardia restrictus to (22S, 25S)-22, 26-epimino-1,4-cholestadien-3-one and (22R, 25S)-22,26-epimino-1,4-cholestadien-3-one respectively. (20S,22ξ,25ξ)-26-Acetylamino-5α-furostan-3β-ol was dehydrogenated to (20S,22ξ,25ξ)-26-acetylamino-1,4-furostadien-3-one with low yield only; whereas (25ξ)-26-amino-5α-cholestane-3β, 16β,22ξ-triol was acetylated to (25ξ)-26-acetylamino-5α-cholestane-3β, 16β, 22ξ-triol. No degradation of the 1, 4-diene-3-ones by Nocardia restrictus was observed.

N-methylated products of the Solanum steroidal alkaloids tomatidine and solasodine

The Solanum steroidal alkaloids tomatidine and solasodine contain a spiro-ring junction with azaketal functionality. The conversion of the natural products to their N-methylated derivatives involves intermediates in which the formerly spiro-carbon (C-22) is sp2 hybridized, and therefore the stereochemical information at this centre is lost. 13C N.m.r. analysis is used to show that methylation of tomatidine (22S,25S) results in one (22S,25S) product, while the same treatment on solasodine (22R,25R) affords two isomers that can equilibrate in solution, with 22R,25R(major) and 22S,25R(minor) stereochemistry. The 13C n.m.r.-derived conformations of the products suggest an explanation for these results.