Enzyme relaxation in the reaction catalyzed by triosephosphate isomerase: detection and kinetic characterization of two unliganded forms of the enzyme.

Triosephosphate isomerase has been shown to exist in two unliganded forms, one of which binds and isomerizes (R)-glyceraldehyde 3-phosphate and the other of which binds and isomerizes dihydroxyacetone 3-phosphate. The tracer perturbation method of Britton demonstrates the kinetic significance of the interconversion of these two enzyme forms at high substrate concentrations and yields a rate constant of about 10(6) s-1 for the interconversion. Although the molecular nature of the two forms of unliganded enzyme is not defined by these experiments, a shuffling of protons among active site residues, or a protein conformational change, or both, may be involved. This study, coupled with the known rate constants for the substrate-handling steps of triosephosphate isomerase catalysis, completes the kinetic characterization of the catalytic cycle for this enzyme.