초록
<P><B>Abstract</B></P><P><SMALL>l</SMALL>‐lysine is an important amino acid in animals and humans and NADPH is a vital cofactor for maximizing the efficiency of <SMALL>l</SMALL>‐lysine fermentation. Dihydrodipicolinate reductase (DHDPR), an NAD(P)H‐dependent enzyme, shows a variance in nucleotide‐cofactor affinity in bacteria. In this study, we rationally engineered <I>Corynebacterium glutamicum</I> DHDPR (<I>Cg</I>DHDPR) to switch its nucleotide‐cofactor specificity resulting in an increase in final titer (from 82.6 to 117.3 g L<SUP>−1</SUP>), carbon yield (from 0.35 to 0.44 g [g glucose]<SUP>−1</SUP>) and productivity (from 2.07 to 2.93 g L<SUP>−1</SUP> hr<SUP>−1</SUP>) of <SMALL>l</SMALL>‐lysine in JL‐6 Δ<I>dapB::Ec</I>‐<I>dapB</I><SUP>C115G,G116C</SUP> in fed‐batch fermentation. To do this, we comparatively analyzed the characteristics of <I>Cg</I>DHDPR and <I>Escherichia coli</I> DHDPR (<I>Ec</I>DHDPR), indicating that hetero‐expression of NADH‐dependent <I>Ec</I>DHDPR increased <SMALL>l</SMALL>‐lysine production. Subsequently, we rationally modified the conserved structure of cofactor‐binding motif, and results indicated that introducing the mutation K11A or R13A in <I>Cg</I>DHDPR and introducing the mutation R16A or R39A in <I>Ec</I>DHDPR modifies the nucleotide‐cofactor affinity of DHDPR. Lastly, the effects of these mutated DHDPRs on <SMALL>l</SMALL>‐lysine production were investigated. The highest increase (26.2%) in <SMALL>l</SMALL>‐lysine production was observed for JL‐6 Δ<I>dapB::Ec</I>‐<I>dapB</I><SUP>C115G,G116C</SUP>, followed by JL‐6 <I>Cg</I>‐<I>dapB</I><SUP>C37G,G38C</SUP> (21.4%) and JL‐6 Δ<I>dapB::Ec</I>‐<I>dapB</I><SUP>C46G,G47C</SUP> (15.2%). This is the first report of a rational modification of DHDPR that enhances the <SMALL>l</SMALL>‐lysine production and yield through the modulation of nucleotide‐cofactor specificity.</P>