Selective production of red azaphilone pigments in a Monascus purpureus mppDEG deletion mutant

The Monascus azaphilone (MAz) pigment is a well-known food colorant that has yellow, orange and red components. The structures of the yellow and orange MAz differ by two hydride reductions, with yellow MAz being the reduced form. Orange MAz can be non-enzymatically converted to red MAz in the presence of amine derivatives. It was previously demonstrated that mppE and mppG are involved in the biosynthesis of yellow and orange MAz, respectively. However, ${\Delta}mppE$ and ${\Delta}mppG$ knockout mutants maintained residual production of yellow and orange MAz, respectively. In this study, we deleted the region encompassing mppD, mppE and mppG in M. purpureus and compared the phenotype of the resulting mutant (${\Delta}mppDEG$) with that of an mppD knockout mutant (${\Delta}mppD$). It was previously reported that the ${\Delta}mppD$ strain retained the ability to produce MAz but at approximately 10% of the level observed in the wildtype strain. A chemical analysis demonstrated that the ${\Delta}mppDEG$ strain was still capable of producing both yellow and orange MAz, suggesting the presence of minor MAz route(s) not involving mppE or mppG. Unexpectedly, the ${\Delta}mppDEG$ strain was observed to accumulate fast-eluting pigments in a reverse phase high-performance liquid chromatography analysis. A LC-MS analysis identified these pigments as ethanolamine derivatives of red MAz, which had been previously identified in an mppE knockout mutant that produces high amounts of orange MAz. Although the underlying mechanism is largely unknown, this study has yielded an M. purpureus strain that selectively accumulates red MAz.