Non-enzymatic glycosylation (or glycation) and inhibition of the pig heart cytosolic aspartate aminotransferase by glyceraldehyde 3-phosphate

Glyceraldehyde 3-phosphate (Glyc3P), a glycolytic intermediate, non-enzymatically glycosylated (or glycated) and inhibited the pig heart cytoplasmic aspartate aminotransferase (cAAT). Glyc3P (5.0 mM) decreased cAAT activity by 47% after 1 min at 23 degrees C. cAAT activity remained unchanged after a 24 h incubation with either glucose 6-phosphate (5.0 mM) or ribose 5-phosphate (5.0 mM). increasing the incubation pH from 6.4 to 7.8 or the incubation temperature from 23 degrees C to 50 degrees C enhanced Glyc3P's inhibitory effect on cAAT activity. Glyc3P (250-500 mu M) decreased the thermal stability of cAAT as evidenced by lowering the T-m or temperature that caused a 50% irreversible loss of cAAT activity (69 degrees C, control, 58.5 degrees C, 500 mu M Glyc3P). Glyc3P decreased cAAT amino group content and increased glycation products, which were measured by adduct formation, fluorescence and protein crosslinking.