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• W2792639104 abstract "Valine, which is one of the branched-chain amino acids (BCAAs) essential for humans, is widely used in animal feed, dietary supplements and pharmaceuticals. At the commercial level, valine is usually produced by bacterial fermentation from glucose. However, valine biosynthesis can also proceed in the yeast Saccharomyces cerevisiae, which is a useful microorganism in fermentation industry. In S. cerevisiae, valine biosynthesis is regulated by valine itself via the feedback inhibition of acetohydroxyacid synthase (AHAS), which consists of two subunits, the catalytic subunit Ilv2 and the regulatory subunit Ilv6. In this study, to improve the valine productivity of yeast cells, we constructed several variants of Ilv6 by introducing amino acid substitutions based on a protein sequence comparison with the AHAS regulatory subunit of E. coli. Among them, we found that the Asn86Ala, Gly89Asp and Asn104Ala variants resulted in approximately 4-fold higher intracellular valine contents compared with those in cells with the wild-type Ilv6. The computational analysis of Ilv6 predicted that Asn86, Gly89 and Asn104 are located in the vicinity of a valine-binding site, suggesting that amino acid substitutions at these positions induce conformational change of the valine-binding site. To test the effects of these variants on AHAS activity, both recombinant Ilv2 and Ilv6 were purified and reconstituted in vitro. The Ilv6 variants were much less sensitive to feedback inhibition by valine than the wild-type Ilv6. Only a portion of the amino acid changes identified in the E. coli AHAS regulatory subunit IlvH enhanced the valine synthesis, suggesting structural and/or functional differences between the S. cerevisiae and E. coli AHAS regulatory subunits. It should also be noted that these amino acid substitutions did not affect the intracellular pools of the other BCAAs, leucine and isoleucine. The approach described here could be a practical method for the development of industrial yeast strains with high-level production of valine or isobutanol." @default.
• W2792639104 created "2018-03-29" @default.
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• W2792639104 date "2018-03-01" @default.
• W2792639104 modified "2023-10-16" @default.
• W2792639104 title "High-level production of valine by expression of the feedback inhibition-insensitive acetohydroxyacid synthase in Saccharomyces cerevisiae" @default.
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• W2792639104 doi "https://doi.org/10.1016/j.ymben.2018.02.011" @default.
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