Identification of the target amino acids in the site-specific inactivation of triose phosphate isomerase by ferrate anion.
Ferrate anion, an analog of orthophosphate anion, very rapidly inactivates triose phosphate isomerase from chicken muscle. The inactivation can be prevented by the presence of competitive inhibitors. Of the 247 amino acids known to be present in each of the identical monomers of this dimeric enzyme, Trp-168, located in the active site pocket, as well as Trp-191 and His-248 are destroyed. The partial loss of Tyr-164 also occurs. Trp-168 is known from published crystallographic studies to be located in the active site cavity harboring Glu-165. The gamma-carboxylate group of Glu-165 is believed to serve as the nucleophile which catalyzes the isomerization. Tyr-164, Glu-165, and Trp-168 are known to be conserved in all of the triose phosphate isomerases which have been sequenced, including those obtained from mammals, chicken, fish, yeast, and bacteria. It is suggested that the chemical modification of Trp-168 alters its shape and hydrophobic character in a manner that adversely affects the conformation of the active site cavity. When the chicken enzyme is treated with ferrate in the presence of the competitive inhibitor phosphoglycolate, only His-248 is destroyed. Thus, His-248, which is the COOH-terminal amino acid, cannot be essential for activity. This observation is consistent with the knowledge that it is not invariant in the enzyme from various species.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001665:Binding Sites; D002238:Carbohydrate Epimerases / Q000037:antagonists & inhibitors*; D002645:Chickens; D007501:Iron / Q000494:pharmacology*; D007700:Kinetics; D009132:Muscles / Q000201:enzymology; D010446:Peptide Fragments / Q000032:analysis; D011485:Protein Binding; D013056:Spectrophotometry, Ultraviolet; D014305:Triose-Phosphate Isomerase / Q000037:antagonists & inhibitors*
D000596:Amino Acids; D010446:Peptide Fragments; C017311:ferrate ion; D007501:Iron; D002238:Carbohydrate Epimerases; D014305:Triose-Phosphate Isomerase