Within the mitochondrial matrix, pyruvate reacts with coenzyme A, forming CO₂, acetyl CoA, and NADH (Figure 12-14, stage II, left). This reaction, catalyzed by pyruvate dehydrogenase, is highly exergonic (ΔG°′ = −8.0 kcal/mol) and essentially irreversible. Influx of calcium from the MAM into the mitochondrion increases the activity of pyruvate dehydrogenase, driving the formation of acetyl CoA.

Acetyl CoA is a molecule consisting of a two-carbon acetyl group covalently linked to a longer molecule known as coenzyme A (CoA) (Figure 12-15). It plays a central role in the oxidation of pyruvate, fatty acids, and amino acids. In addition, it is an intermediate in numerous biosynthetic reactions, including the transfer of an acetyl group to histone and many other mammalian proteins and the synthesis of lipids such as cholesterol. In respiring mitochondria, however, the two-carbon acetyl group of acetyl CoA is almost always oxidized to CO₂ via the citric acid cycle. Note that the two carbons in the acetyl group come from pyruvate; the third carbon of pyruvate is released as carbon dioxide.