Add time:07/13/2019         Source:sciencedirect.com

The anthraquinones endocrocin and emodin are synthesized by a special class of type I NR-PKSs and a discrete MβL-TE. In this work, we first reconstituted a biosynthetic pathway of endocrocin and emodin in S. cerevisiae by combining enzymes from different sources. We functionally characterized a TE-less NR-PKS (SlACAS) and a MβL-TE (SlTE) from S. lycopersici as well as four orthologous MβL-TEs. SlACAS was coexpressed with different MβL-TEs in S. cerevisiae. SlACAS generated the highest amount of endocrocin when coupled with HyTE, the yield was 115.6% higher than that with the native SlTE. To accumulate more emodin, seven decarboxylases with high homology to HyDC were identified and introduced into the biosynthetic pathway. Among these orthologs, AfDC exhibited the highest catalytic activity and the conversion rate reached 98.6%. A double-point mutant acetyl-CoA carboxylase, ACC1S659A, S1157A, was further introduced to increase the production of malonyl-CoA as a precursor of these anthraquinones. The production of endocrocin (233.6 ± 20.3 mg/L) and emodin (253.2 ± 21.7 mg/L) then dramatically increased. We also optimized the carbon source in the medium and conducted fed-batch fermentation with the engineered strains. The titers of endocrocin and emodin obtained were 661.2 ± 50.5 mg/L and 528.4 ± 62.7 mg/L, respectively, which are higher than previously reported. In this work, by screening a small library of orthologous biosynthetic bricks, an efficient biosynthetic pathway of endocrocin and emodin was first created in S. cerevisiae. This study provides a novel metabolic engineering approach for optimization of the production of desired molecules.

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