The active centre of triose phosphate isomerase

Abstract

The molecular weight and amino acid composition of triose phosphate isomerase have been determined. The molecular weight (43000) is lower and the molecular activity (500000) higher than those of most other glycolytic enzymes. Reaction with iodoacetate (studied with radioactive reagent) takes place in two phases: in the first phase, at pH6.3, cysteine and methionine groups react and enzymic activity is unimpaired; in the second phase, histidine reacts and enzymic activity is lost. Photo-oxidation leads to inactivation, with loss of cysteine, of histidine and of tryptophan, but little loss of tyrosine. The mechanism postulated for the action of the enzyme demands the intervention of a group functioning as a base, and the results obtained are consistent with histidine's being the basic group in the active centre.

The Isolation and Crystallization of Yeast and Rabbit Liver Triose Phosphate Isomerase and a Comparative Characterization with the Rabbit Muscle Enzyme

“…Following incubation an aliquot of the solution was diluted and assayed for enzyme activity remaining. Controls were handled in the same manner other than for the omission of the inhibitors No significant differences in our comparison of rabbit muscle and liver triose phosphate isomerase either in total amino acids, tryptophan content (Table S), thiol content ( Table 9) or N-terminal Previous studies of Burton and Waley have shown that loss of activity with the rabbit muscle enzyme through alkylation and photooxidation paralleled most closely the destruction of histidine residues, suggesting that an imidazole group functioned in the active enzyme [5]. The high resistance of yeast triose phosphate isomerase to inactivation by these means consequently points to major structural difference concerning its conformation and/or catalytic site.…”

“…The analysis of rabbit muscle triose phosphate isomerase by Burton and Waley [5] are included after having been adjusted by us for a Fig.7. Effect of p H on triose phosphate isomerase activity.…”

“…This has led to an alternate proposal that the multiple froms may represent conformers [4], a term first advanced by Kaplan to designate enzymes which are believed to have the same primary structure and molecular weight but which exist in varying conformations [27]. The possibility that the subforms may represent an inadvertent modification of the enzyme during the isolation procedure [5] does not appear likely since Scopes has observed multiple bands of triose phosphate isomerase isomerase activity in fresh raw extracts [IS].…”

“…high molecular activity with the conversion of 500 000 moles of glyceraldehyde-3-phosphate per minute per mole of enzyme [ 5 ] . Histidine [5] and lysine [B] are thought to function in the active site.…”

“…high molecular activity with the conversion of 500 000 moles of glyceraldehyde-3-phosphate per minute per mole of enzyme [ 5 ] . Histidine [5] and lysine [B] are thought to function in the active site. Isotopic studies suggest a catalytic mechanism involving a one proton exchange between the substrate and medium with the formation of an intermediary enolate ion bound to the enzyme 171.…”