Bile Salt 3α- and 12α-Hydroxysteroid Dehydrogenases from Eubacterium lentum and Related Organisms

Abstract

Thirty-two strains ofEubacterium lentum and phenotypically similar anaerobic gram-positive bacilli were screened for intracellular bile salt 3a-and 12a-hydroxysteroid dehydrogenase (HSDHase) activities. These organisms were categorized into four groups: (A) those containing 12a-HSDHase only (10 strains), (B) those containing 3a-and 12a-HSDHase (13 strains), (C) those containing 3a-HSDHase only (2 strains), and (D) those devoid of any measurable HSDHase activity (7 strains). Of the respective four groups, 9/10, 13/13, 0/2, and 0/7 were like the neotype strain of E. lentum (ATCC 25559) in that they produced H2S in a triple sugar iron agar butt, reduced nitrate to nitrite, and weakly decomposed hydrogen peroxide. The other strains were variable for nitrate reduction and activity on hydrogen peroxide, but all the organisms in the first three categories (with one exception) were H2S producers (triple sugar iron agar butt) and all (with one exception) were designated E. lentum, whereas the organisms of category B were non-H2S producers (triple sugar iron agar butt). Five of these seven were not stimulated by arginine and are designated "phenotypically similar organisms." Thin-layer chromatography of extracted spent bacterial medium of four representative strains from each group grown in the presence of cholate revealed the presence of (A) .12-oxo product, (B) 12-oxo and 3-oxo products, (C) 3-oxo product, and (D) the absence of any of these products. The 12a-HSDHase of category B organisms was unstable unless 10'-M dithioerythritol was added to the buffer. With the exception of 3 out of 32 strains, there was a positive correlation between the production of measurable amounts of 12a-HSDHase and H2S production. Growth curves and the effect of arginine on growth and the production of 3a-and 12a-HSDHase were examined in representative strains of categories A, B, and C.

Targeted Synthesis and Characterization of a Gene Cluster Encoding NAD(P)H-Dependent 3α-, 3β-, and 12α-Hydroxysteroid Dehydrogenases from Eggerthella CAG:298, a Gut Metagenomic Sequence

“…The high K m and low turnover number and catalytic efficiency of CDD59473 in the reductive direction (Table 2), which partially agree with values previously reported for E. lenta DSM 2243 T 3␤-HSDH, may further explain why iso-CDCA does not predominate in culture (13). Taken together, this three-gene cluster accounts for three of the four HSDH activities reported in E. lenta strains (13,23,24) (Fig. 4).…”

“…Identification of a gene cluster predicted to encode multiple HSDHs in a metagenomic sequence. 12␣-HSDHs (EC 1.1.1.176) have been reported in gut bacteria such as Clostridium perfringens (27), Clostridium leptum (28), E. lenta (23), and Clostridium sp. strain ATCC 29733/VPI C48-50 (15).…”

“…Eggerthella lenta (formerly Eubacterium lentum) is a Gram-positive, non-sporeforming, nonmotile, obligate anaerobic bacterium whose growth is stimulated by arginine (17,18). E. lenta is a particularly important participant in the gut sterolbiome, various strains of which have the potential to encode enzymes that metabolize cardiac glycosides (19,20) and 21-dehydroxylate corticosteroids (21), hydrolyze conjugated bile acids (22), and oxidize bile acid hydroxyl groups (23,24). E. lenta strains have previously been grouped according to their pattern of hydroxysteroid dehydrogenase (HSDH) activities, with some strains having no HSDH activity (group V) and others expressing 3␣-HSDH, 3␤-HSDH, 7␣-HSDH, and 12␣-HSDH (group III) (24).…”

Transformation of Bile Acids by Mixed Microbial Cultures from Human Feces and Bile Acid Transforming Activities of Isolated Bacterial Strains

“…The bacteriological features of the HA-24 group conform to the description of Eubact~rium lentum by Moore et al (21), and the reduction of nitrate and the enhancement of growth by arginine and citrulline satisfy the two positive criteria proposed by Macdonald et al (13) for the identification of E. lentum. Regarding the transformation of bile acids by this species, Midtvedt and Norman (17) reported the presence of 7a-dehydrogenation as well as 3a-epimerization in cultures of three strains of the genus Eubacterium including one of E. lentum, but Macdonald et al (13,14) could not demonstrate either activity in cell-free preparations from the neotype strain and many local isolates of E. lentum, although 3a-and 12a-hydroxysteroid dehydrogenases were found to be common among these strains. The results of the cultural experiments with the present E. lentum strains are consistent with the findings of Midtvedt and Norman, and substantial levels of this species in the human intestinal tract (2, 6) afford a satisfactory explanation for the constant occurrence of 3a-epimerization in fecal cultures and also the frequent appearance of these 3,B-epimers in vivo.…”

Microbiological evaluation of periapical infections in Egypt

“…Eubacterium was frequently present in cultures containing BPB, which suggests sotne fortn of cotntnetisalism or synergistn between the 2 categories. Strains of E. lenttttn have been reported to produce steroid-degrading enzymes (20,34), which can be of importance in providing BPB with growth-protnoting naphthoquinones (32,38,40). Sperry & Wilkins (34) indicate that cells of i;.…”