Method for increasing yield of L-arginine by knocking out Flavin reductases

Abstract

The invention discloses a method for increasing the yield of L-arginine by knocking out flavin reductases, and belongs to the technical field of amino acid production by microbial fermentation. Genes frd1 and frd2 for encoding hypothetic NADPH-dependent FMN reductase in Corynebacterium crenatum SDNN403 are over-expressed in E. coli BL21 and are purified to form target proteins Frd181 and Frd188, and functions of the target proteins are identified to obtain a result showing that the proteins Frd181 and Frd188 both are NAD(P)H-dependent flavin reductases producing H2O2. By taking a genome of the Corynebacterium crenatum SDNN403 as a template, frd1 and frd2 gene deletion fragments are obtained by overlap extension PCR; connecting pK18mobsacB to obtain knockout plasmids pK18mobsacB-Δfrd1 and pK18mobsacB-Δfrd2; carrying out electric shock to transform the Corynebacterium crenatum SDNN403; and carrying out secondary screening to obtain recombinant strains 403Δfrd1 and 403Δfrd2. Found by flask shaking fermentation, the yield of L-arginine is obviously increased by knocking out the genes frd1 and frd2.

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