title: Initial step in the catabolism of cholesterol by Mycobacterium smegmatis mc2 155.
abstract: The first step in the catabolism of cholesterol, i.e. the transformation of cholesterol into cholestenone, has been investigated in Mycobacterium smegmatis. In silico analysis identified the MSMEG_1604 gene encoding a putative protein similar to the ChoD cholesterol oxidase of M. tuberculosis H37Rv (Rv3409c) and the MSMEG_5228 gene coding for a protein similar to the NAD(P)-dependent cholesterol dehydrogenase/isomerase of Nocardia sp. The expression of the MSMEG_5228 gene was inducible by cholesterol whereas the expression of MSMEG_1604 gene was constitutive. When both genes were expressed in Escherichia coli only the MSMEG_5228 protein was active on cholesterol. The function of ChoD-like MSMEG_1604 protein remains to be elucidated, but it does not appear to play a critical role in the mineralization of cholesterol as a MSMEG_1604(-) mutant was not affected in the production of cholestenone. However, a MSMEG_5228(-) mutant showed a drastic reduction in the synthesis of cholestenone. The finding that this mutant was still able to grow in cholesterol, allowed us to demonstrate that the cholesterol-inducible MSMEG_5233 gene encodes an additional cholesterol dehydrogenase/isomerase similar to the AcmA dehydrogenase of Sterolibacterium denitrificans. The observation that the double MSMEG_5228-5233(-) mutant was able to grow in cholesterol suggests that in addition to these enzymes other dehydrogenase/isomerases can also catalyse the first reaction of the cholesterol degradation pathway in M. smegmatis, which is not the limiting step of the process.
authors.affiliation: Environmental Biology Department, Centro de Investigaciones Biológicas, CSIC, Ramiro de Maeztu, 9, 28040 Madrid, Spain.
authors.forename: I
authors.initials: I
authors.lastname: Uhía
authors.forename: B
authors.initials: B
authors.lastname: Galán
authors.forename: V
authors.initials: V
authors.lastname: Morales
authors.forename: J L
authors.initials: JL
authors.lastname: García
mesh_terms: D000595:Amino Acid Sequence; D002783:Cholestenones / Q000378:metabolism; D002784:Cholesterol / Q000378:metabolism*; D004252:DNA Mutational Analysis; D004269:DNA, Bacterial / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D017353:Gene Deletion; D007535:Isomerases / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009154:Mutation; D020102:Mycobacterium smegmatis / Q000201:enzymology* / Q000235:genetics; D010088:Oxidoreductases / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
chemical_list: D002783:Cholestenones; D004269:DNA, Bacterial; D002784:Cholesterol; D010088:Oxidoreductases; C019884:cholesterol dehydrogenase; D007535:Isomerases