Enzyme and Microbial Technology, Vol.111, 57-62, 2018
Essential role of amino acid position 71 in substrate preference by meso-diaminopimelate dehydrogenase from Symbiobacterium thermophilum IAM14863
meso-Diaminopimelate dehydrogenase (meso-DAPDH) catalyzes the reversible oxidative deamination of the d-configuration of meso-2,6diaminopimelate (meso-DAP) and is thought to have substrate specificity toward meso-DAP. The discovery of the meso-DAPDH from Symbiobacterium thermophilum IAM14863 (StDAPDH) revealed meso-DAPDH members with broad substrate specificity. In order to elucidate the substrate-preference mechanism of StDAPDH, it is necessary to identify the key residues related to this mechanism. Our previous work suggested that the non-active-site R71 of StDAPDH was related to substrate preference. Here, we report the key roles of the non-active site on the catalysis of StDAPDH. In order to explore the mechanism through which non active-site R71 only affected the amination activity of StDAPDH, we performed molecular dynamic simulations and investigated the functional role of R71 in the type II meso-DAPDH StDAPDH. Site-directed mutagenesis with the allelic site A69 of CgDAPDH as a target proved that when replaced by Arg at position 71 of StDAPDH, the CgA69R mutant showed higher catalytic efficiencies toward a series of 2-keto acids, ranging from 1.2-to 1.5 fold. These findings provide some guidelines for improving our understanding of the broad substrate specificity of StDAPDH.