Metabolic engineering of Mucor circinelloides for zeaxanthin production

Mucor circinelloides is a β-carotene producing zygomycete amenable to metabolic engineering using molecular tools. The crtS gene of the heterobasidiomycetous yeast Xanthophyllomyces dendrorhous encodes the enzymatic activities β-carotene hydroxylase and ketolase, allowing this yeast to produce the xanthophyll called astaxanthin. Here we describe the fermentation of X. dendrorhous in astaxanthin producing conditions to purify mRNA for the cloning of the cDNA from the crtS gene by RT-PCR. Further construction of an expression plasmid and transformation of M. circinelloides protoplasts allow the heterologous expression of the crtS cDNA in M. circinelloides to obtain β-cryptoxanthin and zeaxanthin overproducing transformants. These two xanthophylls are hydroxylated compounds from β-carotene. These results show that the crtS gene is involved in the conversion of β-carotene into xanthophylls, being potentially useful to engineer carotenoid pathways.