Acyl-CoA dehydrogenase

Class of enzymes that catalyze fatty acid β oxidation in mitochondria
Acyl CoA dehydrogenases (ACADs) are a class of enzymes that function to catalyze the initial step in each cycle of fatty acid β oxidation in the mitochondria of cells. Their action results in the introduction of a trans double bond between C2 (α) and C3 (β) of the acyl CoA thioester substrate. Flavin adenine dinucleotide (FAD) is a required co factor in addition to the presence of an active site glutamate in order for the enzyme to function. The following reaction is the oxidation of the fatty acid by FAD to afford an α,β unsaturated fatty acid thioester of Coenzyme A:

ACADs can be categorized into three distinct groups based on their specificity for short , medium , or long chain fatty acid acyl CoA substrates. While different dehydrogenases target fatty acids of varying chain length, all types of ACADs are mechanistically similar. Differences in the enzyme occur based on the location of the active site along the amino acid sequence. nematodes, fungi, and bacteria. Five of these nine classes are involved in fatty acid β oxidation (SCAD, MCAD, LCAD, VLCAD, and VLCAD2), and the other four are involved in branched chain amino acid metabolism (i3VD, i2VD, GD, and iBD). Most acyl CoA dehydrogenases are α4 homotetramers, and in two cases (for very long chain fatty acid substrates) they are α2 homodimers. An additional class of acyl CoA dehydrogenase was discovered that catalyzes α,β unsaturation reactions with steroid CoA thioesters in certain types of bacteria. This class of ACAD was demonstrated to form α2β2 heterotetramers, rather than the usual α4 homotetramer, a protein architecture that evolved in order to accommodate a much larger steroid CoA substrate. ACADs are classified as .

The medium chain acyl CoA dehydrogenase (MCAD) is the best known structure of all ACADs, and is the most commonly deficient enzyme within the class that leads to metabolic disorders in animals. The interface between the two monomers of a single dimer of an ACAD contains the FAD binding sites and has extensive bonding interactions. In contrast, the interface between the two dimers has fewer interactions. There are a total of 4 active sites within the tetramer, each of which contains a single FAD molecule and an acyl CoA substrate binding site. This gives a total of four FAD molecules and four acyl CoA substrate binding sites per enzyme. FAD is bound between the three domains of the monomer, where only the nucleotide portion is accessible. FAD binding contributes significantly to overall enzyme stability. The acyl CoA substrate is bound completely within each monomer of the enzyme. The active site is lined with the residues F252, T255, V259, T96, T99, A100, L103, Y375, Y375, and E376. The area of interest within the substrate becomes wedged between Glu 376 and FAD, lining up the molecules into an ideal position for the reaction. A novel ACAD enzyme architecture in some species of steroid utilizing bacteria (Actinomycetota and Pseudomonadota) was discovered, and is involved in the utilization of ubiquitous steroid substrates like cholesterol by pathogenic organisms like Mycobacterium tuberculosis. Genetically, the structure is encoded by two separate genes (open reading frames) that form an obligate α2β2 heterotetramic complex. The structure was most likely the result of an evolutionary event that caused gene duplication and partial loss of function, since half of the FAD cofactor binding residues are in each gene, and only make a complete binding site when expressed together as a complex. This probably allowed for the substrate binding site to open up considerably to accommodate much larger polycyclic CoA substrates, rather than fatty acids of varying chain lengths.

The acyl CoA dehydrogenase mechanism proceeds through an E2 elimination. This elimination is initiated by a glutamate residue, which, while necessary for the mechanism, is not conserved. In humans the most common naturally occurring mutation in MCAD is located at amino acid residue Lys 304. This can lead to the symptoms of the deficiency listed above.