Improved Bioconversion of 15-Fluoro-6-deoxyerythronolide B to 15-Fluoro-erythromycin A by Overexpression of the eryK Gene in Saccharopolyspora erythraea

The bioconversion of a 6-deoxyerythronolide B analogue to the corresponding erythromycin A analogue (R-EryA) by a Saccharopolyspora erythraea mutant lacking the ketosynthase in the first polyketide synthase module was significantly improved by changing fluxes at a key branch point affecting the erythromycin congener distribution. This was achieved by integrating an additional copy of the eryK gene into the chromosome under control of the eryAIp promoter. Real-time PCR analysis of RNA confirmed higher expression of eryK in the resulting strain, S. erythraea K301-105B, compared to its parent. In shake flasks, K301-105B produced less of the shunt product 15-fluoro-erythromycin B (15F-EryB), suggesting a shift in congener distribution toward the desired product, 15-fluoro-erythromycin A (15F-EryA). In bioreactor studies, K301-105B produced 1.3 g/L of 15F-EryA with 75-80% molar yield on fed precursor, compared with 0.9 g/L 15F-EryA with 50-55% molar yield on fed precursor by the parent strain. At higher precursor feed rates, K301-105B produced 3.5 g/L of 15F-EryA while maintaining 75-80% molar yield on fed precursor.