Acute acrylonitrile toxicity: Inhibition of glyceraldehyde-3-phosphate dehydrogenase activity in vitro

Acrylonitrile (AN) is an organic compound produced in large quantities by the chemical industry. It is used as a monomer in the production of various polymers. Acrylonitrile is an acute toxin. The long-term goal of this research is to identify the mechanism responsible for the acute lethality of acrylonitrile. Several mechanisms have been proposed to explain the toxicity of acrylonitrile: AN depletion of glutathione (GSH) with subsequent oxidative tissue damage, metabolism of AN by P450 followed by the release of the toxic metabolite cyanide (CN) and covalent binding of AN to tissue proteins with subsequent loss of protein function.; This study addresses this last mechanism. We focused on proteins with known reactive cysteine residues. One very attractive candidate was glyceraldehyde-3-phosphate dehydrogenase (GAPDH), a key enzyme in glycolysis. Incubation of rabbit muscle GAPDH with AN, in vitro, led to irreversible inhibition of enzyme activity that followed pseudo-first order kinetics. The observed first order rate constant was proportional to the AN concentration over the range of 50–400 μM, with no evidence of saturation. From this data the second order rate constant for inhibition of 3.8 M−1 sec−1 at 25°C was calculated. At 37°C, this rate constant increased two-fold. These data are consistent with the hypothesis that inhibition of GAPDH may be the “other” mechanism, besides cyanide toxicity, to account for the acute toxicity of AN. Understanding the mechanism of AN toxicity has an importance that goes far beyond that of AN alone. Many exogenous or endogenous compounds (e.g. reactive oxygen species) can react with cysteine residues in proteins, possibly altering protein function. Understanding the mechanism of toxicity of AN will help design better treatments for AN intoxication, as well as for compounds with a similar mechanism of toxicity.