Biosynthesis of mono-acylated mannosylerythritol lipid in an acyltransferase gene-disrupted mutant of Pseudozyma tsukubaensis

The basidiomycetous yeast genus Pseudozyma produce large amounts of mannosylerythritol lipids (MELs), which are biosurfactants. A few Pseudozyma strains produce mono-acylated MEL as a minor compound using excess glucose as the sole carbon source. Mono-acylated MEL shows higher hydrophilicity than di-acylated MEL and has great potential for aqueous applications. Recently, the gene cluster involved in the MEL biosynthesis pathway was identified in yeast. Here, we generated an acyltransferase ( PtMAC2 ) deletion strain of P. tsukubaensis 1E5 with uracil auxotrophy as a selectable marker. A PtURA5 -mutant with a frameshift mutation in PtURA5 was generated as a uracil auxotroph of strain 1E5 by ultraviolet irradiation on plate medium containing 5-fluoro-orotic acid (5-FOA). In the mutant, PtMAC2 was replaced with a PtURA5 cassette containing the 5′ untranslated region (UTR) (2000 bp) and 3′ UTR (2000 bp) of PtMAC2 by homologous recombination, yielding strain Δ PtMAC2 . Based on TLC and NMR analysis, we found that Δ PtMAC2 accumulates MEL acylated at the C-2′ position of the mannose moiety. These results indicate that PtMAC2p catalyzes acylation at the C-3′ position of the mannose of MEL.