Alda‐1 modulates the kinetic properties of mitochondrial aldehyde dehydrogenase (ALDH2)

• Published in: The FEBS journal Vol. 283; no. 19; pp. 3637 - 3650
• Main Authors: Belmont-Diaz, Javier A, Yoval-Sanchez, Belem, Calleja-Castaneda, Luis F, Pardo Vazquez, Juan P, Rodriguez-Zavala, Jose S
• Format: Journal Article
• Language: English
• Published: England Blackwell Publishing Ltd 01-10-2016
• Subjects: Alda‐1; Aldehyde Dehydrogenase, Mitochondrial - chemistry; Aldehyde Dehydrogenase, Mitochondrial - drug effects; Aldehyde Dehydrogenase, Mitochondrial - metabolism; Aldehydes; ALDH2; Benzamides - chemistry; Benzamides - pharmacology; Benzodioxoles - chemistry; Benzodioxoles - pharmacology; Binding sites; cardioprotective drug; Cysteine - chemistry; Dehydrogenases; Enzyme Activators - chemistry; Enzyme Activators - pharmacology; enzyme kinetics; Kinetics; Mitochondria; rate‐limiting step; cardioprotective drug; ALDH2; rate-limiting step; enzyme kinetics; Alda-1
• ISSN: 1742-464X; 1742-4658
• DOI: 10.1111/febs.13833

Mitochondrial aldehyde dehydrogenase (ALDH2) has been proposed as a key enzyme in cardioprotection during ischemia–reperfusion processes. This proposal led to the search for activators of ALDH2 with the aim to develop cardioprotective drugs. Alda‐1 was the first activator of ALDH2 identified and its cardioprotective effect has been extensively proven in vivo; however, the mechanism of activation is not fully understood. A crystallographic study showed that Alda‐1 binds to the entrance of the aldehyde‐binding site; therefore, Alda‐1 should in essence be an inhibitor. In the present study, kinetic experiments were performed to characterize the effect of Alda‐1 on the properties of ALDH2 (kinetic parameters, determination of the rate‐limiting step, reactivity of the catalytic cysteine) and on the kinetic mechanism (type of kinetics, sequence of substrates entering, and products release). The results showed that Alda‐1 dramatically modifies the properties of ALDH2, the Km for NAD+ decreased by 2.4‐fold, and the catalytic efficiency increased 4.4‐fold; however, the Km for the aldehyde increased 8.6‐fold, thus, diminishing the catalytic efficiency. The alterations in these parameters resulted in a complex behavior, where Alda‐1 acts as inhibitor at low concentrations of aldehyde and as an activator at high concentrations. Additionally, the binding of Alda‐1 to ALDH2 made the deacylation less limiting and diminished the pKa of the catalytic cysteine. Finally, NADH inhibition patterns indicated that Alda‐1 induced a change in the sequence of substrates entry and products release, in agreement with the proposal of both substrates entering ALDH2 by the NAD+ entrance site.
Activator of human ALDH2 Alda‐1 protects cell integrity from diverse stress conditions; however, the activation mechanism is not fully understood. Kinetic characterization showed that binding of Alda‐1 to ALDH2 activates the catalytic cysteine reducing its pKa, which induces the switching of order of substrates binding and products release, additionally, Alda‐1 increases the rate of the limiting step of the reaction.