Roles of D-lactate dehydrogenases in the anaerobic growth of Shewanella oneidensis MR-1 on sugars

Abstract

Shewanella oneidensis MR-1 is a facultative anaerobe that respires using a variety of electron acceptors. Although this organism is incapable of fermentative growth in the absence of electron acceptors, its genome encodes LdhA (a putative fermentative NADH-dependent D-lactate dehydrogenase [D-LDH]) and Dld (a respiratory quinone-dependent D-LDH). However, the physiological roles of LdhA in MR-1 are unclear. Here, we examined the activity, transcriptional regulation, and traits of deletion mutants to gain insight into the roles of LdhA in the anaerobic growth of MR-1. Analyses of D-LDH activity in MR-1 and the ldhA deletion mutant confirmed that LdhA functions as an NADH-dependent D-LDH that catalyzes the reduction of pyruvate to D-lactate. In vivo and in vitro assays revealed that ldhA expression was positively regulated by the cyclic-AMP receptor protein, a global transcription factor that regulates anaerobic respiratory pathways in MR-1, suggesting that LdhA functions in coordination with anaerobic respiration. Notably, we found that a deletion mutant of all four NADH dehydrogenases (NDHs) in MR-1 (ΔNDH mutant) retained the ability to grow on N-acetylglucosamine under fumarate-respiring conditions, while an additional deletion of ldhA or dld deprived the ΔNDH mutant of this growth ability. These results indicate that LdhA-Dld serves as a bypass of NDH in electron transfer from NADH to quinones. Our findings suggest that the LdhA-Dld system manages intracellular redox balance by utilizing D-lactate as a temporal electron sink under electron acceptor-limited conditions.