Improvement of FK506 Production in the High-Yielding Strain Streptomyces sp. RM7011 by Engineering the Supply of Allylmalonyl-CoA Through a Combination of Genetic and Chemical Approach

Abstract

FK506, a widely used immunosuppressant, is a 23-membered polyketide macrolide that is produced by several Streptomyces species. FK506 high-yielding strain Streptomyces sp. RM7011 was developed from the discovered Streptomyces sp. KCCM 11116P by random mutagenesis in our previous study. The results of transcript expression analysis showed that the transcription levels of tcsA, B, C, and D were increased in Streptomyces sp. RM7011 by 2.1-, 3.1-, 3.3-, and 4.1- fold, respectively, compared with Streptomyces sp. KCCM 11116P. The overexpression of tcsABCD genes in Streptomyces sp. RM7011 gave rise to approximately 2.5-fold (238.1 μg/ml) increase in the level of FK506 production compared with that of Streptomyces sp. RM7011. When vinyl pentanoate was added into the culture broth of Streptomyces sp. RM7011, the level of FK506 production was approximately 2.2-fold (207.7 μg/ml) higher than that of the unsupplemented fermentation. Furthermore, supplementing the culture broth of Streptomyces sp. RM7011 expressing tcsABCD genes with vinyl pentanoate resulted in an additional 1.7-fold improvement in the FK506 titer (498.1 μg/ml) compared with that observed under nonsupplemented condition. Overall, the level of FK506 production was increased approximately 5.2-fold by engineering the supply of allylmalonyl-CoA in the high-yielding strain Streptomyces sp. RM7011, using a combination of overexpressing tcsABCD genes and adding vinyl pentanoate, as compared with Streptomyces sp. RM7011 (95.3 μg/ml). Moreover, among the three precursors analyzed, pentanoate was the most effective precursor, supporting the highest titer of FK506 in the FK506 high-yielding strain Streptomyces sp. RM7011.