<P> Some variability also exists at the previous step--the conversion of 2 - oxoglutarate to succinyl - CoA . While most organisms utilize the ubiquitous NAD - dependent 2 - oxoglutarate dehydrogenase, some bacteria utilize a ferredoxin - dependent 2 - oxoglutarate synthase (EC 1.2. 7.3). Other organisms, including obligately autotrophic and methanotrophic bacteria and archaea, bypass succinyl - CoA entirely, and convert 2 - oxoglutarate to succinate via succinate semialdehyde, using EC 4.1. 1.71, 2 - oxoglutarate decarboxylase, and EC 1.2. 1.79, succinate - semialdehyde dehydrogenase . </P> <P> The regulation of the citric acid cycle is largely determined by product inhibition and substrate availability . If the cycle were permitted to run unchecked, large amounts of metabolic energy could be wasted in overproduction of reduced coenzyme such as NADH and ATP . The major eventual substrate of the cycle is ADP which gets converted to ATP . A reduced amount of ADP causes accumulation of precursor NADH which in turn can inhibit a number of enzymes . NADH, a product of all dehydrogenases in the citric acid cycle with the exception of succinate dehydrogenase, inhibits pyruvate dehydrogenase, isocitrate dehydrogenase, α - ketoglutarate dehydrogenase, and also citrate synthase . Acetyl - coA inhibits pyruvate dehydrogenase, while succinyl - CoA inhibits alpha - ketoglutarate dehydrogenase and citrate synthase . When tested in vitro with TCA enzymes, ATP inhibits citrate synthase and α - ketoglutarate dehydrogenase; however, ATP levels do not change more than 10% in vivo between rest and vigorous exercise . There is no known allosteric mechanism that can account for large changes in reaction rate from an allosteric effector whose concentration changes less than 10% . </P> <P> Calcium is also used as a regulator in the citric acid cycle . Calcium levels in the mitochondrial matrix can reach up to the tens of micromolar levels during cellular activation . It activates pyruvate dehydrogenase phosphatase which in turn activates the pyruvate dehydrogenase complex . Calcium also activates isocitrate dehydrogenase and α - ketoglutarate dehydrogenase . This increases the reaction rate of many of the steps in the cycle, and therefore increases flux throughout the pathway . </P> <P> Citrate is used for feedback inhibition, as it inhibits phosphofructokinase, an enzyme involved in glycolysis that catalyses formation of fructose 1, 6 - bisphosphate, a precursor of pyruvate . This prevents a constant high rate of flux when there is an accumulation of citrate and a decrease in substrate for the enzyme . </P>

Which of the following is the net reaction for two turns of the citric acid cycle