title: Alda-1 modulates the kinetic properties of mitochondrial aldehyde dehydrogenase (ALDH2).
abstract: 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 Kmfor NAD+decreased by 2.4-fold, and the catalytic efficiency increased 4.4-fold; however, the Kmfor 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 pKaof 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.
authors: Departamento de Bioquímica, Instituto Nacional de Cardiología, México D.F, México.
authors: Javier A
authors: JA
authors: Belmont-Díaz
authors: Departamento de Bioquímica, Instituto Nacional de Cardiología, México D.F, México.
authors: Belem
authors: B
authors: Yoval-Sánchez
authors: Departamento de Bioquímica, Instituto Nacional de Cardiología, México D.F, México.
authors: Luis F
authors: LF
authors: Calleja-Castañeda
authors: Departamento de Bioquímica, Facultad de Medicina, Universidad Nacional Autónoma de México, México D.F, México.
authors: Juan P
authors: JP
authors: Pardo Vázquez
authors: Departamento de Bioquímica, Instituto Nacional de Cardiología, México D.F, México. rodjos@cardiologia.org.mx, rzjs@yahoo.com.
authors: José S
authors: JS
authors: Rodríguez-Zavala
mesh_terms: D000071396:Aldehyde Dehydrogenase, Mitochondrial / Q000737:chemistry / Q000187:drug effects* / Q000378:metabolism; D001549:Benzamides / Q000737:chemistry / Q000494:pharmacology*; D052117:Benzodioxoles / Q000737:chemistry / Q000494:pharmacology*; D003545:Cysteine / Q000737:chemistry; D020536:Enzyme Activators / Q000737:chemistry / Q000494:pharmacology*; D007700:Kinetics
chemical_list: D001549:Benzamides; D052117:Benzodioxoles; D020536:Enzyme Activators; C531525:N-(1,3-benzodioxol-5-ylmethyl)-2,6-dichlorobenzamide; C096127:ALDH2 protein, human; D000071396:Aldehyde Dehydrogenase, Mitochondrial; D003545:Cysteine
keywords: ALDH2; Alda-1; cardioprotective drug; enzyme kinetics; rate-limiting step