Studies Of Acyltransferases And Pathway-Specific Regulators Involved In FK506Biosynthesis

FK506(also named as tacrolimus), a23-membered ring macrolide antibiotic, is mainly used as immunosuppressive, antibacterial, and anticancer medicines with high value in market. FK506is synthesized by a polyketide synthase-nonribosomal peptide synthetase (PKS-NRPS) hybrid. Here we studied the biosynthetic pathway of FK506in an industrial FK506producing strain Streptomyces tsukubaensis YN06.We characterized AT10FkbA, an essential enzyme involved in FK506biosynthesis by bioinformatics and biochemical assay. Our results showed the function of AT10FkbA is to transfer malonyl (M) from malonyl coenzyme A (M-CoA) into itself to form M-O-AT10FkbA intermediate and then to transfer M from M-O-AT10FkbA into holo-ACP10FkbA to form M-S-ACP10FkbA.In the first step self-acylation reaction, AT10FkbA has broad substrate specificity for M-CoA and methylmalonyl coenzyme A (MM-CoA). In the second step transacylation reaction, AT10FkbA has strict substrate specificity for M-CoA. We also in vivo characterized pathways-specific regulatory factor Tcs2. Our results showed that Tcs2may play a positive regulating role in FK506biosynthesis, although its target was not found in the FK506biosynthetic gene cluster by EMSA. Finally, FK506production was increased by about20%by overexpression of tcs2in S.tsukubaensis YN06.