Pyruvate ferredoxin oxidoreductase from the hyperthermophilic archaeon, Pyrococcus furiosus , functions as a CoA-dependent pyruvate decarboxylase

Abstract

Pyruvate ferredoxin oxidoreductase (POR) has been previously purified from the hyperthermophilic archaeon, Pyrococcus furiosus, an organism that grows optimally at 100°C by fermenting carbohydrates and peptides. The enzyme contains thiamine pyrophosphate and catalyzes the oxidative decarboxylation of pyruvate to acetyl-CoA and CO 2 and reduces P. furiosus ferredoxin. Here we show that this enzyme also catalyzes the formation of acetaldehyde from pyruvate in a CoA-dependent reaction. Desulfocoenzyme A substituted for CoA showing that the cofactor plays a structural rather than a catalytic role. Ferredoxin was not necessary for the pyruvate decarboxylase activity of POR, nor did it inhibit acetaldehyde production. The apparent K m values for CoA and pyruvate were 0.11 mM and 1.1 mM, respectively, and the optimal temperature for acetaldehyde formation was above 90°C. These data are comparable to those previously determined for the pyruvate oxidation reaction of POR. At 80°C (pH 8.0), the apparent V m value for pyruvate decarboxylation was about 40% of the apparent V m value for pyruvate oxidation rate (using P. furiosus ferredoxin as the electron acceptor). Tentative catalytic mechanisms for these two reactions are presented. In addition to POR, three other 2-keto acid ferredoxin oxidoreductases are involved in peptide fermentation by hyperthermophilic archaea. It is proposed that the various aldehydes produced by these oxidoreductases in vivo are used by two aldehyde-utilizing enzymes, alcohol dehydrogenase and aldehyde ferredoxin oxidoreductase, the physiological roles of which were previously unknown.

Single gene insertion drives bioalcohol production by a thermophilic archaeon

“…2B), acetyl-CoA is not likely to be the source of this acetaldehyde for ethanol production. Acetaldehyde could arise in P. furiosus from the decarboxylation of pyruvate, which was previously shown to be a significant side reaction of POR (19). Alternatively, it could arise by the reduction of acetate by P. furiosus aldehyde Fd oxidoreductase (AOR).…”

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Both Forward and Reverse TCA Cycles Operate in Green Sulfur Bacteria

“…The amount of acetate was determined by a coupled acetylCoA synthetase/citrate synthase/malate dehydrogenase assay following the formation of NADH (17).…”