Pyruvate is a key precursor metabolite for the biosynthesis of building blocks under both aerobic and anaerobic conditions. It gives, upon dissimilation, acetyl-CoA, which is also an important biosynthetic precursor metabolite. Under anaerobic conditions and in the absence of external electron acceptors, pyruvate is also the most prominent intermediate metabolite for the synthesis of most fermentative products, including lactate, formate, ethanol, acetate, carbon dioxide, and hydrogen.
The conversion of pyruvate to lactate is achieved via a fermentative pathway catalyzed by the enzyme lactate dehydrogenase. Two different enzymes catalyze the conversion of pyruvate into acetyl-CoA: pyruvate formate lyase (PFL) and pyruvate dehydrogenase (PDH). PFL is known to be active under conditions of very low or no oxygen. On the other hand, it is widely accepted that the main role of PDH is in the aerobic dissimilation of pyruvate. In recent years however, it has been shown that PDH is in fact active under anaerobic and fermentative conditions, although its role is not known. A comparison of the PDH and PFL reactions for the conversion of pyruvate into acetyl-CoA reveals that the reaction catalyzed by PDH produces NADH (reducing power) as opposed to the alternative pathway (catalyzed by PFL), which results in no net generation of reducing equivalents. There is one more enzyme involved in the dissimilation of pyruvate, pyruvate oxidase (PoxB). PoxB catalyses the decarboxylation of pyruvate to acetate and CO2, a reaction that also generates reducing equivalents in the form of flavin adenine dinucleotide. The metabolism of pyruvate via PoxB is less efficient than the route via the PDH; however, the PoxB route is important for wild-type growth efficiency and responsible for a significant amount of pyruvate metabolism under aerobic conditions. However, its metabolic role (if any) under fermentative conditions is still unknown.
The objective of this project is to elucidate the physiological role of the aforementioned pyruvate-dissimilating enzymes during anaerobic fermentation of different carbon sources by E. coli. Using traditional genetic and biochemical approaches in combination with metabolic flux analysis we have discovered that PDH plays an important role in the fermentative metabolism of many carbon sources including glucose. Depending on the carbon source fermented, the role of PDH is related either to the fulfillment of cellular requirement for carbon dioxide and/or the maintenance of redox balance. In both cases, PDH performs these functions in a more carbon, energy, and redox efficient manner than alternative pathways.