Biological activities of an engineered tautomycetin analogue via disruption of tmcR-encoding hydroxylase in Streptomyces sp. CK4412

Tautomycetin (TMC), originally isolated from Streptomyces griseochromogenes, has been reported to possess biological functions including T cell-specific immunosuppressive and anticancer activities through a mechanism of differential inhibition of protein phosphatases such as PP1, PP2A, and SHP2. Independently isolated Streptomyces sp. CK4412 was also reported to produce a structurally identical TMC compound. Previously, we isolated and characterized the entire TMC biosynthetic gene cluster from Streptomyces sp. CK4412. In silico database comparison revealed a 1,359-bp tmcR as a putative bacterial Cytochrome P450 hydroxylase gene in the TMC biosynthetic gene cluster. Through targeted gene disruption and complementation, the tmcR mutant was confirmed to produce a C5-deoxy-TMC, the same analogue produced by the S. griseochromogenes ttnI mutant, implying that TmcR behaves as a regiospecific C5-oxygenase in the TMC biosynthetic pathway in Streptomyces sp. CK4412. In particular, the C5-deoxy-TMC from the tmcR mutant exhibited 3.2-fold higher inhibition activity toward SHP2 with significantly reduced inhibition activities toward PP1, and human Vero and lung cancer cells. These results suggested that C5 regiospecific modification of the TMC polyketide moiety may result in a drug development target for use in preferentially enhancing immunosuppressive activity while minimizing its undesirable biological activities.