Investigation of the pre-steady-state kinetics of fructose bisphosphatase by employment of an indicator method.
The pre-steady-state kinetics for the hydrolysis of fructose 1,6-bisphosphate by rabbit liver fructose bis-phosphatase have been investigated by stopped-flow kinetics utilizing an acid-base indicator method that permits the continuous monitoring of the inorganic phosphate product. The reaction sequence is characterized by two successive first-order steps followed by establishment of the steady-state rate. The first exponential process results from a conformational change in the protein that is dye sensitive owing to a perturbation of an acidic residue on the protein. A second process reflects the rapid initial turnover of all four subunits of the enzyme with the concomitant release of inorganic phosphate followed by the rate-limiting step of the catalytic cycle. This latter step may involve a product release (fructose 6-phosphate) or a second conformational change. The catalytic cycle ends with decay of the enzyme to its initial unreactive resting state.
D000818:Animals; D002384:Catalysis; D003201:Computers; D006597:Fructose-Bisphosphatase / Q000378:metabolism*; D005635:Fructosediphosphates / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D046911:Macromolecular Substances; D010637:Phenolsulfonphthalein; D011487:Protein Conformation; D011817:Rabbits
D005635:Fructosediphosphates; D046911:Macromolecular Substances; D006597:Fructose-Bisphosphatase; D010637:Phenolsulfonphthalein