Biology subjects

Feller, F. M.

Publications and source records attributed to Feller, F. M..

2 recordsLinked to original sources

bioRxiv2021

Comparative analysis of bile-salt degradation in Sphingobium sp. strain Chol11 and P. stutzeri Chol1 reveals functional diversity of β-proteobacterial steroid degradation enzymes and suggests a novel reaction sequence for side-chain degradation involving a hydroxylation step

The reaction sequence for aerobic degradation of bile salts by environmental bacteria resembles degradation of other steroid compounds. Recent findings show that bacteria belonging to the Sphingomonadaceae use a pathway variant for bile-salt degradation. This study addresses this so-called {Delta}4,6 -variant by comparative analysis of unknown degradation steps in Sphingobium sp. strain Chol11 with known reactions found in Pseudomonas stutzeri Chol1. Investigations with strain Chol11 revealed an essential function of the acyl-CoA dehydrogenase Scd4AB for growth with bile salts. Growth of the scd4AB deletion mutant was restored with a metabolite containing a double bond within the side chain which was produced by the {Delta}22-acyl-CoA dehydrogenase Scd1AB from P. stutzeri Chol1. Expression of scd1AB in the scd4AB deletion mutant fully restored growth with bile salts while expression of scd4AB only enabled constricted growth in P. stutzeri Chol1 scd1A or scd1B deletion mutants. Strain Chol11 {Delta}scd4A accumulated hydroxylated steroid metabolites which were degraded and activated with coenzyme A by the wild type. Activities of five Rieske type monooxygenases of strain Chol11 were screened by heterologous expression and compared to the B-ring cleaving KshABChol1 from P. stutzeri Chol1. Three of the Chol11 enzymes catalyzed B-ring cleavage of only {Delta}4,6 -steroids while KshABChol1 was more versatile. Expression of a fourth KshA homolog, Nov2c228 led to production of metabolites with hydroxylations at an unknown position. These results indicate functional diversity of {beta}-proteobacterial enzymes for bile-salt degradation and suggest a novel side-chain degradation pathway involving an essential ACAD reaction and a steroid hydroxylation step. ImportanceThis study highlights the biochemical diversity of bacterial degradation of steroid compounds in different aspects. First, it further elucidates an unexplored variant in the degradation of bile-salt side chains by Sphingomonads, a group of environmental bacteria that is well-known for their broad metabolic capabilities. Moreover, it adds a so-far unknown hydroxylation of steroids to the reactions Rieske monooxygenases can catalyze with steroids. Additionally, it analyzes the first proteobacterial ketosteroid-9-hydroxylase and shows that this enzyme is able to catalyze side reactions with non-native substrates.

microbiology↗

bioRxiv2021

Proteome, bioinformatic and functional analyses reveal a distinct and conserved metabolic pathway for bile salt degradation in the Sphingomonadaceae

Bile salts are amphiphilic steroids with a C5 carboxylic side chain with digestive functions in vertebrates. Upon excretion, they are degraded by environmental bacteria. Degradation of the bile-salt steroid skeleton resembles the well-studied pathway for other steroids like testosterone, while specific differences occur during side-chain degradation and the initiating transformations of the steroid skeleton. Of the latter, two variants via either {Delta}1,4- or {Delta}4,6-3-ketostructures of the steroid skeleton exist for 7-hydroxy bile salts. While the {Delta}1,4- variant is well-known from many model organisms, the {Delta}4,6-variant involving a 7-hydroxysteroid dehydratase as key enzyme has not been systematically studied. Here, combined proteomic, bioinformatic and functional analyses of the {Delta}4,6-variant in Sphingobium sp. strain Chol11 were performed. They revealed a degradation of the steroid rings similar to the {Delta}1,4-variant except for the elimination of the 7-OH as key difference. In contrast, differential production of the respective proteins revealed a putative gene cluster for side-chain degradation encoding a CoA-ligase, an acyl-CoA dehydrogenase, a Rieske monooxygenase, and an amidase, but lacking most canonical genes known from other steroid-degrading bacteria. Bioinformatic analyses predicted the {Delta}4,6-variant to be widespread among the Sphingomonadaceae, which was verified for three type strains which also have the predicted side-chain degradation cluster. A second amidase in the side-chain degradation gene cluster of strain Chol11 was shown to cleave conjugated bile salts while having low similarity to known bile-salt hydrolases. This study signifies members of the Sphingomonadaceae remarkably well-adapted to the utilization of bile salts via a partially distinct metabolic pathway. ImportanceThis study highlights the biochemical diversity of bacterial degradation of steroid compounds, in particular bile salts. Furthermore, it substantiates and advances knowledge of a variant pathway for degradation of steroids by sphingomonads, a group of environmental bacteria that are well-known for their broad metabolic capabilities. Biodegradation of bile salts is a critical process due to the high input of these compounds from manure into agricultural soils and wastewater treatment plants. In addition, these results may also be relevant for the biotechnological production of bile salts or other steroid compounds with pharmaceutical functions.

microbiology↗