SUMMARY
Mycolic acids are essential components of the mycobacterial cell wall. In this study we show
that a gene encoding a reductase involved in the final step of mycolic acid biosynthesis can be
deleted in Mycobacterium smegmatis without affecting cell viability. Deletion of MSMEG4722
(ortholog of Mycobacterium tuberculosis Rv2509) altered culture characteristics and antibiotic
sensitivity. The ∆MSMEG4722 strain synthesized α-alkyl, β-oxo intermediates of mycolic acids
which were found esterified to cell wall-arabinogalactan. While the precursors could not be
isolated directly due to their inherent instability during base-treatment, their presence was
established by prior reduction of the β-oxo group by sodium borohydride. Interestingly, the
mutant also accumulated unsaturated ketones, similar to tuberculenone from M. tuberculosis,
which were shunt products derived from spontaneous decarboxylation of α-alkyl, β-oxo fatty
acid precursors of mycolic acids.