Oxidative Stress Evokes a Metabolic Adaptation That Favors Increased NADPH Synthesis and Decreased NADH Production in Pseudomonas fluorescens

Abstract

ABSTRACT The fate of all aerobic organisms is dependent on the varying intracellular concentrations of NADH and NADPH. The former is the primary ingredient that fuels ATP production via oxidative phosphorylation, while the latter helps maintain the reductive environment necessary for this process and other cellular activities. In this study we demonstrate a metabolic network promoting NADPH production and limiting NADH synthesis as a consequence of an oxidative insult. The activity and expression of glucose-6-phosphate dehydrogenase, malic enzyme, and NADP + -isocitrate dehydrogenase, the main generators of NADPH, were markedly increased during oxidative challenge. On the other hand, numerous tricarboxylic acid cycle enzymes that supply the bulk of intracellular NADH were significantly downregulated. These metabolic pathways were further modulated by NAD + kinase (NADK) and NADP + phosphatase (NADPase), enzymes known to regulate the levels of NAD + and NADP + . While in menadione-challenged cells, the former enzyme was upregulated, the phosphatase activity was markedly increased in control cells. Thus, NADK and NADPase play a pivotal role in controlling the cross talk between metabolic networks that produce NADH and NADPH and are integral components of the mechanism involved in fending off oxidative stress.