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ChoD Is Important for Lipid Modification in M. smegmatis
Author(s) -
Gao Jin,
Sampson Nicole S.
Publication year - 2008
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.22.1_supplement.611.17
Subject(s) - mycobacterium smegmatis , cholesterol oxidase , biochemistry , chemistry , mycobacterium , enzyme , biology , mycobacterium tuberculosis , bacteria , genetics , tuberculosis , medicine , pathology
New targets are needed to develop antimicrobial therapies to combat M. tuberculosis ( Mtb ) , especially the multi and extremely drug resistant organisms (MDR‐TB and XDR‐TB). Mycobacteria have been reported to oxidize 3β ‐hydroxysterols to 3‐ketosteroids, but the enzyme(s) responsible for this activity have not been identified in mycobacterial species. Rv3409c, from Mycobacteria tuberculosis H37Rv genome is annotated as choD , a putative cholesterol oxidase. The choD gene was heterologously expressed behind an acetamidase promoter with a C‐terminal hexa‐histidine tag in Mycobacterium smegmatis mc 2 155. Using established cholesterol oxidase assays that follow the formation of the by‐product H 2 O 2 or final product cholest‐4‐en‐3‐one, there was no cholesterol oxidizing activity detected. The choD gene is also conserved in M. smegmatis , M. leprae and Rhodococcus equi. The M. smegmatis orthologue has 83% amino acid identity with choD from M. tuberculosis . The M. smegmatis choD transposon mutant Myc11 can still catabolize cholesterol. Moreover, Myc11 has significantly different colony morphology than wild‐type as observed by light microscopy. The total lipids from both M. smegmatis WT and Myc11 were extracted and analyzed by MALDI‐TOF MS and TLC. There are distinct differences between the Myc11 lipid profile and the wild‐type lipid profile. Based on our results, we propose that choD is not a cholesterol oxidase, but rather plays a role in lipid cell wall modification or synthesis.