Structural studies of intersubunit communication in mitochondrial aldehyde dehydrogenase

Mitochondrial aldehyde dehydrogenase, ALDH2, plays a principal role in ethanol metabolism by catalyzing the NAD+-dependent oxidation of acetaldehyde. ALDH2 has also been identified as a bioactivator of nitroglycerin. Evidence for intersubunit communication within this tetrameric enzyme has been uncovered in both the observed half-site reactivity of wild type and the semi-dominance of ALDH2*2, a polymorphism found in 40% of the East Asian population. The glutamate to lysine substitution at residue 487 in ALDH2*2 leads to low activity and is associated with an aversive response to alcohol consumption and reduced efficacy of nitroglycerin in the treatment of angina pectoris.;ALDH2 is considered a dimer of dimers where glutamate 487 is located at the intra-dimer interface and forms hydrogen bonds with arginine 264 of the same subunit and arginine 475 of the adjacent subunit, stabilizing the interface and the coenzyme binding site. For ALDH2*2, the lysine at 487 produces an enzyme that demonstrates an elevated KMNAD+ and decreased kcat. In an attempt to rescue ALDH2*2, arginine 475 was mutated to glutamine (R475Q). Although this modification did not restore wild-type-like activity to ALDH2*2, positive cooperativity for NAD+ binding was unexpectedly induced by this substitution. From these results we hypothesized that the interaction of arginines 264 and 475 and glutamate 487 influence subunit communication in the form of dominance and cooperativity.;X-ray crystallography has been employed to investigate the structural features that underlie both ALDH2*2 dominance and R475Q cooperativity. The apoenzyme ALDH2*2 and R475Q structures demonstrate disorder surrounding residues 246, 487, and 475, with ALDH2*2 being more severely affected. The binding of NAD+ to either enzyme partially alleviates this disorder. These structures show that glutamate 487 links the coenzyme-binding site to the active site through arginines 264 and 475, and in doing so, creates a stable structural scaffold conducive to catalysis. Disruption of this network is communicated across the interface, disturbing both coenzyme-binding sites, leading to impaired activity and altered NAD+-binding properties, the degrees of which are determined by the magnitude of structural perturbations. Thus, mechanisms for both the dominance of the ALDH2*2 allele and the positive cooperativity of the R475Q mutant are revealed.