Biotransformation of lauric acid into 1,12-dodecanedioic acid using CYP52A17 expressed in Saccharomyces cerevisiae and its application in refining coconut factory wastewater

Abstract

1,12-Dodecanedioic acid (DDA), a primary compound and an intermediate precursor for various chemical products, is normally produced by chemical synthesis, which presents potential disadvantages. Instead, the biosynthesis of 1,12-DDA by recombinant (r) microorganisms may offer a viable production route. Cytochrome P450 (CYP) can terminally oxidize fatty acids to ω-hydroxy-fatty acids and further to dicarboxylic acids (ω-oxidation). The wild type (r) CYP52A17LL and its engineered L261S/L490S form, in which the two leucine residues were changed into serine (rCYP52A17SS), were expressed in Pichia pastoris and a strain of Saccharomyces cerevisiae coexpressing the yeast NADPH cytochrome P450 reductase gene. Both organisms produced 12-hydroxydodecanoic acid (HDDA) and 1,12-DDA from lauric acid. In vitro, microsomes containing rCYP52A17SS extracted from the S. cerevisiae strain BY(2R)/pYeDP60-CYP52A17SS were able to convert lauric acid to 12-HDDA quite efficiently. Biotransformation of lauric acid using S. cerevisiae BY(2R)/pYeDP60-CYP52A17SS in culture gave the highest level of 12-HDDA (45.8 μM) at 24 h, which was oxidized to yield 20.8 μM of 1,12-DDA at 72 h. The recombinant S. cerevisiae BY(2R)/pYeDP60-CYP52A17SS, which was initially cultured in YPGE, produced the highest yield of 1,12-DDA from coconut milk wastewater at 24 h. Hence, our designed S. cerevisiae strain BY(2R)/pYeDP60-CYP52A17SS can potentially produce 1,12-DDA for industrial applications.