Acyl-CoA dehydrogenase

Structure

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.

MCAD can bind to a rather broad range of chain-lengths in the acyl-CoA substrate, however studies show that its specificity tends to target octanoyl-CoA (C8-CoA).

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.

Mechanism

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.

The residue appears in a wide range of locations within the different types of the enzyme (it is Glu 376 in MCAD). The glutamate residue deprotonates the pro-R hydrogen of the alpha carbon. Hydrogen bonding of the substrate's carbonyl oxygen to both the 2'-OH of the ribityl side-chain of FAD and to the main chain N-H of the previously mentioned glutamate residue lowers the pKa of this proton, allowing it to be readily removed by glutamate.

As the alpha carbon is being deprotonated, the pro-R hydrogen of the beta carbon leaves as a hydride to FAD in a concerted step. It adds to the Re face of FAD at the N-5 position, and the enzyme holds FAD in place through hydrogen bonding with the pyrimidine portion and hydrophobic interactions with the dimethylbenzene portion. The substrate has now been transformed into an α,β unsaturated thioester.

As FAD picks up the hydride, the carbonyl oxygen adjacent to the N-1 nitrogen becomes negatively charged. These electrons are in resonance with the N-1 nitrogen, distributing and stabilizing the resulting negative charge. The charge is also stabilized by hydrogen bonding between the oxygen and nitrogen of interest and various residues within the enzyme.

Clinical significance

Deficiencies in acyl-CoA dehydrogenases result in decreased ability to oxidize fatty acids, thereby signifying metabolic dysfunction. Medium-chain acyl-CoA dehydrogenase deficiencies (MCADD) are well known and characterized because they occur most commonly among acyl-CoA dehydrogenases, leading to fatty acid oxidation disorders and the potential of life-threatening metabolic diseases. Some symptoms of medium-chain acyl-CoA dehydrogenase deficiency include intolerance to fasting, hypoglycemia, and sudden infant death syndrome. These symptoms are seen as directly connected to the inability to metabolize fats. Intolerance to fasting and hypoglycemia result from the inability to gain energy and make sugar from fat stores, which is how most of humans' excess energy is stored. Also, fatty acids can begin to accumulate in the blood, lowering the blood's pH and causing acidosis.

MCAD is related to / has an association with sudden infant death. Approximately 90% of cases of MCAD are due to a single point mutation where the lysine at position 304 (Lys304) is replaced by a glutamate residue and this prevents the enzyme from properly functioning.

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.

References

• "Molecular graphics images were produced using the UCSF Chimera package from the Resource for Biocomputing, Visualization, and Informatics at the University of California, San Francisco (supported by NIH P41 RR-01081). "

References

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