THE MECHANISM OF REGULATION OF HEXOKINASE - NEW INSIGHTS FROM THE CRYSTAL-STRUCTURE OF RECOMBINANT HUMAN BRAIN HEXOKINASE COMPLEXED WITH GLUCOSE AND GLUCOSE-6-PHOSPHATE

Background: Hexokinase I is the pacemaker of glycolysis in brain tissu e. The type I isozyme exhibits unique regulatory properties in that ph ysiological levels of phosphate relieve potent inhibition by the produ ct, glucose-6-phosphate (Gluc-6-P). The 100 kDa polypeptide chain of h exokinase I consists of a C-terminal (catalytic) domain and an N-termi nal (regulatory) domain. Structures of ligated hexokinase I should pro vide a basis for understanding mechanisms of catalysis and regulation at an atomic level. Results: The complex of human hexokinase I with gl ucose and Gluc-6-P (determined to 2.8 Angstrom resolution) is a dimer with twofold molecular symmetry. The N- and C-terminal domains of one monomer interact with the C-and N-terminal domains, respectively, of t he symmetry-related monomer. The two domains of a monomer are connecte d by a single alpha helix and each have the fold of yeast hexokinase. Salt links between a possible cation-binding loop of the N-terminal do main and a loop of the C-terminal domain may be important to regulatio n. Each domain binds single glucose and Gluc-6-P molecules in proximit y to each other. The 6-phosphoryl group of bound Gluc-6-P at the C-ter minal domain occupies the putative binding site for ATP, whereas the 6 -phosphoryl group at the N-terminal domain may overlap the binding sit e for phosphate. Conclusions: The binding synergism of glucose and Glu c-6-P probably arises out of the mutual stabilization of a common (glu cose-bound) conformation of hexokinase I. Conformational changes in th e N-terminal domain in response to glucose, phosphate, and/or Gluc-6-P may influence the binding of ATP to the C-terminal domain.