NITROPRUSSIDE STIMULATES THE CYSTEINE-SPECIFIC MONO(ADP-RIBOSYLATION)OF GLYCERALDEHYDE-3-PHOSPHATE DEHYDROGENASE FROM HUMAN ERYTHROCYTES

In human erythrocyte membranes incubated with [adenylate-P-32]NAD the 36 kDa protein is predominantly labeled. The labeling is greatly stimu lated by nitroprusside in the presence of dithiothreitol. We have puri fied the 36 kDa protein and identified this modification as cysteine-s pecific mono(ADP-ribosylation) because: (i) labeling occurred only whe n [P-32]NAD was replaced by adenine [U-C-14]NAD, but not by [carbonyl- C-14]NAD; (ii) treatment of the prelabeled protein with snake venom ph osphodiesterase led to releasing 5'[P-32]AMP; (iii) the bond between t he protein and the nucleotide was hydrolyzed by HgCl2, but was resista nt to hydroxylamine. The 36 kDa protein reacted on Western blots with two different monoclonal antibodies (MAbs) against glyceraldehyde-3-ph osphate dehydrogenase (GAPDH) and was immunoprecipitated by both MAbs.