Rat brain hexokinase: Location of the allosteric regulatory site in a structural domain at the N-terminus of the enzyme
| Autor: | Tracy K. White, John E. Wilson |
|---|---|
| Rok vydání: | 1987 |
| Předmět: |
Proteolysis
Allosteric regulation Biophysics Glucose-6-Phosphate Regulatory site Biology Guanidines Peptide Mapping Biochemistry chemistry.chemical_compound Hexokinase medicine Animals Urea Binding site Molecular Biology Guanidine Hexoses chemistry.chemical_classification Binding Sites medicine.diagnostic_test Glucosephosphates Brain Rats Enzyme chemistry Glucose 6-phosphate Allosteric enzyme biology.protein |
| Zdroj: | Archives of Biochemistry and Biophysics. 259:402-411 |
| ISSN: | 0003-9861 |
| Popis: | After denaturation in 0.6 M guanidine hydrochloride, rat brain hexokinase becomes highly susceptible to proteolysis by trypsin. Glucose 6-phosphate (Glc-6-P) and its analog, 1,5-anhydroglucitol 6-phosphate, selectively protect the N-terminal half of the molecule from proteolysis. These compounds do not protect the C-terminal half of the molecule, nor do they protect enzyme activity; the Glc analog, N-acetylglucosamine, does protect the C-terminal domain and catalytic activity, but does not prevent proteolysis of the N-terminal half of the molecule. These results are consistent with previous work [M. Nemat-Gorgani and J. E. Wilson (1986) Arch. Biochem. Biophys. 251, 97-103; D. M. Schirch and J. E. Wilson (1987) Arch. Biochem. Biophys. 254, 385-396] demonstrating that binding sites for both hexose and nucleotide substrates, and thus catalytic function, are associated with a 40-kDa domain located at the C-terminus of the enzyme. They further demonstrate that the binding site for the allosteric effector, Glc-6-P, lies in the N-terminal half of the molecule and is distinct from the catalytic site. Using protection against proteolysis as a reflection of binding, it is shown that the Glc-6-P binding site in the N-terminal region has all the characteristics described for the allosteric effector site on this enzyme in terms of affinity for Glc-6-P, specificity, and synergistic interactions with the hexose binding site in the C-terminal region of the molecule. This disposition of catalytic and regulatory functions in discrete halves of the molecule is consistent with suggestions by several investigators that mammalian hexokinases evolved by a process of duplication and fusion of an ancestral gene coding for a hexokinase similar to the present-day yeast enzyme, with the regulatory site of mammalian hexokinases having evolved from what was originally a catalytic site. |
| Databáze: | OpenAIRE |
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