fragment of m/z 351 corresponding to the α-chain (C22) plus a carbonyl group. In addition, the
detection of fragments of m/z 546, 574 and 602 corresponding to C39, C41 and C43 mono-
unsaturated alkyl chains respectively further substantiated our findings.
DISCUSSION
With the exception of KasB, all enzymes involved in the biosynthesis of mycolic acids in
mycobacterial species are encoded by essential genes (Bhatt et al., 2005; Brown et al., 2007;
Parish et al., 2007; Portevin et al., 2004; Sacco et al., 2007). However, we were able to generate
a viable null mutant of MSMEG4722, the gene that encodes the reductase involved in mycolic
acid motif synthesis in M. smegmatis. This was not entirely surprising because global transposon
mutagenesis screens predicted insertions in Rv2509, the M. tuberculosis homologue of
MSMEG4722, to result in a slow growth phenotype. Indeed, the ∆MSMEG4722 mutant did
exhibit a slow growth rate similar to what was observed in C. glutamicum (Lea-Smith et al.,
2007).
Alkaline hydrolysis of the parental strain M. smegmatis mc2155 released α-, α'- and
epoxy mycolates, as expected, but hydrolysates of the ∆MSMEG4722 mutant had no evidence
for mycolates (Figure 3A). Instead, hydrolysates of the mutant ∆MSMEG4722 showed the
presence of rapidly migrating components (labelled ‘?’, Figure 3A, B). If the ∆MSMEG4722
mutant was accumulating α-alkyl, β-oxo mycolate precursors, alkaline hydrolysis would produce
unstable β-oxo acids, which would lose carbon dioxide to yield long-chain ketones. Using an
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