Article Figures & Data
Figures
- FIG 1
Monod degradation kinetics of androgens by resting cell suspensions of S. denitrificans. The increase of bacterial biomass within 2 h was negligible (<3%). The solid lines represent fitted Monod kinetic curves. The data are from one representative experiment of four individual experiments.
- FIG 2
Denitrifying growth of S. denitrificans DSMZ 13999 on testosterone. (A) Assimilation and mineralization of [4-14C]testosterone in the bacterial culture. Symbols: ▽, ethyl acetate-extractable 14C; ○, assimilated 14C; □, trapped 14CO2. (B) Nitrate consumption and testosterone utilization in the same bacterial culture. Symbols: ●, bacterial growth; ■, residual total testosterone; ▲, nitrate consumption. The data shown are the means ± SE of three experimental measurements.
- FIG 3
UPLC-HRMS analysis of a 13C-labeled intermediate produced by the denitrifying S. denitrificans. (A) UPLC chromatogram of the ethyl acetate extract. (B) ESI mass spectra of the C17 intermediate. The predicted elemental composition was calculated using MassLynx mass spectrometry software (Waters). (C) Expected chemical structure. Critical carbon atoms in this compound are numbered according to the steroidal carbon numbering system. ∗, 13C atom.
- FIG 4
Structural elucidation of 6α-hydroxyandrost-4-en-3,17-dione produced by S. denitrificans. (A) APCI mass spectrum. (B) Key heteronuclear multiple-bond correlation (HMBC) spectrum.
- FIG 5
Time course of intermediate production and androgenic activity of a denitrifying S. denitrificans culture. The A420 of the negative control, DMSO (1%, vol/vol), was set to zero.
- FIG 6
Response of the yeast androgen bioassay to the individual intermediates. The results are from one representative of four individual experiments.
- FIG 7
Testosterone catabolism by S. denitrificans proceeds via the 2,3-seco pathway regardless of oxygen conditions. (A) Testosterone and nitrate consumption in the S. denitrificans cultures. Symbols: ●, testosterone remaining in the anaerobic culture; ■, testosterone in the microaerobic culture; ▲, testosterone in the aerobic culture; ○, nitrate consumption under anaerobic conditions; □, nitrate consumption under microaerobic conditions; △, nitrate consumption under aerobic conditions. (B) 2,3-SAOA production in three S. denitrificans cultures. Representative data from one of three independent experiments are shown.
- FIG 8
(A) Proposed pathway of anoxic testosterone degradation by the denitrifying bacteria. Note that the 6-hydroxysteroid is produced by S. denitrificans DSMZ 13999 only when 3-mercaptopropionate is present. ∗, intermediates detected in S. denitrificans cultures (this study). †, intermediates detected in Sdo. denitrificans DSMZ 18526 (39, 41). (B) The established retro-aldol reaction involved in the 9,10-seco pathway for the oxic cholesterol catabolism (56).
Additional Files
Supplemental material
Files in this Data Supplement:
- Supplemental file 1 -
Phylogenetic analysis of Sterolibacterium strains (Fig. S1), UPLC-APCI-HRMS analysis of the ethyl acetate extract of S. denitrificans (Fig. S2), NMR spectra of the HPLC-purified 6α-hydroxyandrost-4-en-3,17-dione (Fig. S3), and UPLC-ESI-HRMS analyses of the ethyl acetate extracts of Comamonas testosteroni and S. denitrificans grown aerobically on testosterone (Fig. S4).
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- Supplemental file 1 -
