ROUTES OF FLAVODOXIN AND FERREDOXIN REDUCTION IN ESCHERICHIA-COLI - COA-ACYLATING PYRUVATE - FLAVODOXIN AND NADPH - FLAVODOXIN OXIDOREDUCTASES PARTICIPATING IN THE ACTIVATION OF PYRUVATE FORMATE-LYASE

Abstract

Flavodoxin and ferredoxin become reduced in E. coli cells by oxidoreductase reactions which use pyruvate and NADPH as electron donor substrates. The 2 enzymes, which are minor proteins of this organism, were measured through the reduced flavodoxin-dependent activation of pyruvate formate-lyase. The NADPH-dependent enzyme, obtained homogeneously through Procion-red affinity chromatography, was identified as the flavoprotein component R described previously by Fujii and Huennekens. The pyruvate-dependent enzyme was identified as CoA-acetylating pyruvate:flavodoxin (ferredoxin) oxidoreductase. Its catalytic properties in the forward, reverse and the 14CO2-pyruvate exchange reaction are reported. The dihydro form of flavodoxin was characterized as the particular species involved in the activation of pyruvate formate-lyase. The activation process still occurs with 70% of maximal efficiency when the ratio [NADPH]/([NADP] + [NADPH]) is fixed at the intracellular anabolic reduction charge value of 0.45, in conjunction with the NADPH-dependent enzyme. The [2Fe-2S] ferredoxin, though being readily used as electron acceptor of both oxidoreductases and having a redox potential similar to flavodoxin, proved incompetent in mediating the activation of pyruvate formate-lyase.