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. 2022 Nov 4;9(1):116.
doi: 10.1186/s40643-022-00602-7.

Bioproduction of testosterone from phytosterol by Mycolicibacterium neoaurum strains: "one-pot", two modes

Daria N Tekucheva  1 , Vera M Nikolayeva  2 , Mikhail V Karpov  2 , Tatiana A Timakova  2 , Andrey V Shutov  2 , Marina V Donova  2
Affiliations

Bioproduction of testosterone from phytosterol by Mycolicibacterium neoaurum strains: "one-pot", two modes

Daria N Tekucheva et al. Bioresour Bioprocess. .

Abstract

The main male hormone, testosterone is obtained from cheap and readily available phytosterol using the strains of Mycolicibacterium neoaurum VKM Ac-1815D, or Ac-1816D. During the first "oxidative" stage, phytosterol (5-10 g/L) was aerobically converted by Ac-1815D, or Ac-1816D to form 17-ketoandrostanes: androstenedione, or androstadienedione, respectively. At the same bioreactor, the 17-ketoandrostanes were further transformed to testosterone due to the presence of 17β-hydroxysteroid dehydrogenase activity in the strains ("reductive" mode). The conditions favorable for "oxidative" and "reductive" stages have been revealed to increase the final testosterone yield. Glucose supplement and microaerophilic conditions during the "reductive" mode ensured increased testosterone production by mycolicibacteria cells. Both strains effectively produced testosterone from phytosterol, but highest ever reported testosterone yield was achieved using M. neoaurum VKM Ac-1815D: 4.59 g/l testosterone was reached from 10 g/l phytosterol thus corresponding to the molar yield of over 66%. The results contribute to the knowledge on phytosterol bioconversion by mycolicibacteria, and are of significance for one-pot testosterone bioproduction from phytosterol bypassing the intermediate isolation of the 17-ketoandrostanes.

Keywords: Mycolicibacterium neoaurum; 17β-hydroxysteroid dehydrogenase; Phytosterol; Testosterone; “One-pot” bioproduction.

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Conflict of interest statement

The authors declare that they have no competing interests.

Figures

Fig. 1
Fig. 1
Scheme of C19 steroid compounds transformation: red arrows—hydrogenation/dehydrogenation at C1(2). Blue arrows—reduction/oxidation at C17
Fig. 2
Fig. 2
Influence of media composition on phytosterol1 biotransformation by Mycolicibacterium neoaurum VKM Ac-1815D under “oxidative” mode. 1—Medium M1 + 10 g/l glucose; 2—medium M2 + 10 g/l glucose; 3—medium M2 + mCD + 10 g/l glucose; 4—medium M2 + mCD + 20 g/l glucose. 1Initial phytosterol load was 10 g/l. The molar ratio of phytosterol to mCD was 1:0.3. Medium M2 is the medium M1 supplemented with the soy bean flour
Fig. 3
Fig. 3
Effect of aeration modes and their combination on phytosterol transformation by Mycolicibacterium neoaurum VKM Ac-1815D1. A Transformation was carried out only under the “oxidative” mode; B change of the transformation mode from “oxidative” to “reductive” was conducted at 120 h (vertical line). Red graphs—glucose was added daily throughout transformation period (24–196 h); blue graphs—glucose was not added daily; green graphs—glucose was added throughout “reductive” mode (144–196 h). 1The initial phytosterol load was 5 g/l. The molar ratio of phytosterol to mCD was 1:0.3. Glucose (10 g/l) was added to the medium M2 at the inoculation moment (0 h) in all cases and additionally as mentioned in legend
Fig. 4
Fig. 4
Effect of mCD content on phytosterol transformation by Mycolicibacterium neoaurum VKM Ac-1815D1. A On the process duration and phytosterol transformation efficiency2, B on the steroids yields. 1The transformation was performed in M2 with initial phytosterol load of 10 g/l. At the inoculation moment (0 h) 10 g/l of glucose and further 5 g/l daily was added. The duration of “reductive” mode was 72 h for all phytosterol to mCD molar ratios. The duration of “oxidative” mode was varied. 2The efficiency of phytosterol transformation was calculated as total molar concentration of all steroidal products divided by the initial molar phytosterol concentration, %
Fig. 5
Fig. 5
Influence of glucose supplement manner on phytosterol1 bioconversion by Mycolicibacterium neoaurum VKM Ac-1816D under the “oxidative” mode. Green—single glucose addition (10 g/l), orange—daily glucose feeding. 110 g/l of phytosterol was used; the molar ratio of phytosterol to methylated β-cyclodextrin (mCD) was 1:0.8
Fig. 6
Fig. 6
Biotransformation of 10(A) or 5(B) g/l phytosterol into testosterone by Mycolicibacterium neoaurum VKM Ac-1816D under two regimens1. 1the transformation was performed in M2 medium under “oxidative” and “reductive” modes successively with the glucose addition at the inoculation moment (10 g/l) and daily glucose feeding (5 g/l). Molar phytosterol to mCD ratio was 1:0.8
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