title: Kinetic Cooperativity in Human Pancreatic Glucokinase Originates from Millisecond Dynamics of the Small Domain.
abstract: The hallmark of glucokinase (GCK), which catalyzes the phosphorylation of glucose during glycolysis, is its kinetic cooperativity, whose understanding at atomic detail has remained open since its discovery over 40 years ago. Herein, by using kinetic CPMG NMR spectroscopic data for 17 isoleucine side chains distributed over all parts of GCK, we show that the origin of kinetic cooperativity is rooted in intramolecular protein dynamics. Residues of glucose-free GCK located in the small domain displayed distinct exchange behavior involving multiple conformers that are substantially populated (p>17 %) with a kex  value of 509±51 s(-1) , whereas in the glucose-bound form these exchange processes were quenched. This exchange behavior directly competes with the enzymatic turnover rate at physiological glucose concentrations, thereby generating the sigmoidal rate dependence that defines kinetic cooperativity.
authors.affiliation: Department of Chemistry and Biochemistry, The Ohio State University, 100 West 18th Avenue, Columbus, OH 43210 (USA).
authors.forename: Mioara
authors.initials: M
authors.lastname: Larion
authors.affiliation: Department of Chemistry and Biochemistry, The Ohio State University, 100 West 18th Avenue, Columbus, OH 43210 (USA).
authors.forename: Alexandar L
authors.initials: AL
authors.lastname: Hansen
authors.affiliation: National High Magnetic Field Laboratory, Tallahassee, FL 32306 (USA).
authors.forename: Fengli
authors.initials: F
authors.lastname: Zhang
authors.affiliation: Department of Chemistry and Biochemistry, The Ohio State University, 100 West 18th Avenue, Columbus, OH 43210 (USA).
authors.forename: Lei
authors.initials: L
authors.lastname: Bruschweiler-Li
authors.affiliation: Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892-0520 (USA).
authors.forename: Vitali
authors.initials: V
authors.lastname: Tugarinov
authors.affiliation: Department of Chemistry and Biochemistry, Florida State University, Tallahassee, FL 32306 (USA).
authors.forename: Brian G
authors.initials: BG
authors.lastname: Miller
authors.affiliation: Department of Chemistry and Biochemistry, The Ohio State University, 100 West 18th Avenue, Columbus, OH 43210 (USA). bruschweiler.1@osu.edu.
authors.forename: Rafael
authors.initials: R
authors.lastname: Brüschweiler
mesh_terms: D002384:Catalysis; D005941:Glucokinase / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D008958:Models, Molecular; D010179:Pancreas / Q000378:metabolism*; D010766:Phosphorylation
chemical_list: D005941:Glucokinase
keywords: NMR spectroscopy; allostery; enzyme catalysis; kinetic cooperativity; time-resolved conformational dynamics