Catalysis of nitrite generation from nitroglycerin by glyceraldehyde-3-phosphate dehydrogenase (GAPDH)

Abstract

Vascular relaxation to nitroglycerin (glyceryl trinitrate; GTN) requires its bioactivation by mechanisms that remain controversial. We report here that glyceraldehyde-3-phosphate dehydrogenase (GAPDH) catalyzes the release of nitrite from GTN. In assays containing dithiothreitol (DTT) and NAD(+), the GTN reductase activity of purified GAPDH produces nitrite and 1,2-GDN as the major products. A vmax of 2.6nmolmin(-)(1)mg(-)(1) was measured for nitrite production by GAPDH from rabbit muscle and a GTN KM of 1.2mM. Reductive denitration of GTN in the absence of DTT results in dose- and time-dependent inhibition of GAPDH dehydrogenase activity. Disulfiram, a thiol-modifying drug, inhibits both the dehydrogenase and GTN reductase activity of GAPDH, while DTT or tris(2-carboxyethyl)phosphine reverse the GTN-induced inhibition. Incubation of intact human erythrocytes or hemolysates with 2mM GTN for 60min results in 50% inhibition of GAPDH's dehydrogenase activity, indicating that GTN is taken up by these cells and that the dehydrogenase is a target of GTN. Thus, erythrocyte GAPDH may contribute to GTN bioactivation.

Keywords: ALDH2; Bioactivation; DTPA; Erythrocytes; G3P; GAPDH; GDN; GMN; GTN; NAD; NaPPi; Nitrite; Nitroglycerin; TCEP; aldehyde dehydrogenase-2; diethylenetriaminepentaacetic acid; glyceraldehyde-3-phosphate; glyceraldehyde-3-phosphate dehydrogenase; glyceryl dinitrate; glyceryl mononitrate; glyceryl trinitrate (nitroglycerin); heGAPDH; human erythrocyte glyceraldehyde-3-phosphate dehydrogenase; rabbit muscle glyceraldehyde-3-phosphate dehydrogenase; rmGAPDH; sodium pyrophosphate; tris(2-carboxyethyl)phosphine.