ChoG is the main inducible extracellular cholesterol oxidase of Rhodococcus sp. strain CECT3014.
Cholesterol catabolism has been reported in different bacteria and particularly in several Rhodococcus species, but the genetic of this complex pathway is not yet very well defined. In this work we report the isolation and sequencing of a 9.8 kb DNA fragment of Rhodococcus sp. strain CECT3014, a bacterial strain that we here identify as a Rhodococcus erythropolis strain. In this DNA fragment we found several ORF that are probably involved in steroid catabolism, and choG, a gene encoding a putative cholesterol oxidase whose functional characterization we here report. ChoG protein is a class II cholesterol oxidase with all the structural features of the enzymes of this group. The disruption of the choG gene does not alter the ability of strain CECT3014 cells to grow on cholesterol, but it abolishes the production of extracellular cholesterol oxidase. This later effect is reverted when the mutant cells are transformed with a plasmid expressing choG. We conclude that choG is the gene responsible for the inducible extracellular cholesterol oxidase activity of strain CECT3014. This activity distributes between the cellular membrane and the culture supernatant in a way that suggests it is produced by the same ChoG protein that occurs in two different locations. RT-PCR transcript analysis showed a dual scheme of choG expression: a low constitutive independent transcription, plus a cholesterol induced transcription of choG into a polycistronic kstD-hsd4B-choG mRNA.
Fernández de Las Heras
Laura
L
Departamento de Bioquímica y Biología Molecular I, Universidad Complutense de Madrid, 28040 Madrid, Spain.
Mascaraque
Victoria
V
García Fernández
Esther
E
Navarro-Llorens
Juana María
JM
Perera
Julián
J
Drzyzga
Oliver
O
D002784:Cholesterol / Q000378:metabolism; D002789:Cholesterol Oxidase / Q000235:genetics* / Q000378:metabolism*; D016000:Cluster Analysis; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D020869:Gene Expression Profiling; D055786:Gene Knockout Techniques; D005816:Genetic Complementation Test; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D016366:Open Reading Frames; D010802:Phylogeny; D012240:Rhodococcus / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D017422:Sequence Analysis, DNA; D017385:Sequence Homology
D004269:DNA, Bacterial; D002784:Cholesterol; D002789:Cholesterol Oxidase