Identification of active-site residues of sheep liver serine hydroxymethyltransferase.
Chemical modification of amino acid residues with phenylglyoxal, N-ethylmaleimide and diethyl pyrocarbonate indicated that at least one residue each of arginine, cysteine and histidine were essential for the activity of sheep liver serine hydroxymethyltransferase. The second-order rate constants for inactivation were calculated to be 0.016 mM-1 X min-1 for phenylglyoxal, 0.52 mM-1 X min-1 for N-ethylmaleimide and 0.06 mM-1 X min-1 for diethyl pyrocarbonate. Different rates of modification of these residues in the presence and in the absence of substrates and the cofactor pyridoxal 5'-phosphate as well as the spectra of the modified protein suggested that these residues might occur at the active site of the enzyme.
D000818:Animals; D001120:Arginine / Q000032:analysis; D001665:Binding Sites; D003545:Cysteine / Q000032:analysis; D004047:Diethyl Pyrocarbonate / Q000494:pharmacology; D005033:Ethylmaleimide / Q000494:pharmacology; D012696:Glycine Hydroxymethyltransferase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D006639:Histidine / Q000032:analysis; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D010658:Phenylglyoxal / Q000494:pharmacology; D012756:Sheep; D013053:Spectrophotometry; D014166:Transferases / Q000378:metabolism*
D006639:Histidine; D001120:Arginine; D014166:Transferases; D012696:Glycine Hydroxymethyltransferase; D003545:Cysteine; D004047:Diethyl Pyrocarbonate; D010658:Phenylglyoxal; D005033:Ethylmaleimide