Abstract
Microbial steroid metabolism represents an underappreciated extension of the vertebrate endocrine system, with growing evidence that host-associated microbes contribute to the diversity and bioavailability of sex steroids within human tissues. Emerging studies have linked microbial androgen metabolism to urinary microbiome composition and to resistance to androgen deprivation therapy (ADT) in prostate cancer. While microbial pathways capable of converting steroid precursors such as cortisol to androgens, via the steroid-17,20-desmolase pathway, such as DesG-mediated interconversion of androstenedione to testosterone have been reported, the diversity of enzymes mediating downstream androgen interconversion remains incompletely defined. Here, we investigate the androgen-forming capabilities of anaerobic bacteria from the male genitourinary microbiome, focusing on NADPH-dependent 17β-hydroxysteroid dehydrogenases (17β-HSDHs) that catalyze interconversion of androstenedione and testosterone. We isolated androgen-forming bacterial strains from human male urine and identified a previously uncharacterized 17β-HSDH encoded by Peptoniphilus obesi , demonstrated that this enzyme catalyzes the NADPH-dependent reduction of androstenedione to testosterone and the reverse oxidation reaction. Sequence similarity searches further identified a homologous 17β-HSDH in Anaerococcus , which was synthesized and functionally validated, revealing conserved activity despite low sequence identity to the previously characterized urinary tract enzyme DesG. The enzymes were found to have broad substrate specificity for C19 and C18 17keto- and 17β-hydroxysteroids. Together, these findings expand the known diversity of microbial 17β-HSDHs and identify previously unrecognized androgen-forming activities within the genitourinary microbiome. IMPORTANCE: Microbial steroid-transforming pathways may provide a mechanism by which commensal anaerobes contribute to androgen availability in the genitourinary tract. By identifying novel 17β-hydroxysteroid dehydrogenases from Peptoniphilus and Anaerococcus , genera repeatedly associated with prostate cancer, this study provides mechanistic insight into how microbial steroid metabolism may influence hormone-driven disease.