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(The FASEB Journal. 2008;22:lb334)
© 2008 FASEB
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lb334

Glucose modulation of glucokinase activation by small molecules

Erik C. Ralph1, Jim Thomson1, Jonathan Almaden2 and Shaoxian Sun1

1 BiochemPharm, Pfizer, San Diego, CA
2 Biology, UCSD, San Diego, CA

ABSTRACT

Small molecule activators of glucokinase (GK) were used in kinetic and equilibrium binding studies to probe the biochemical basis for their allosteric effects. These small molecules decreased the glucose K0.5 (~1 mM vs. ~8 mM) and the Hill coefficient (1.2 vs. 1.7), and lowered the kcat to 62%–95% of the control activity. They relieved GK's inhibition from GKRP in a glucose-dependent manner and exhibited a competitive relationship with respect to GKRP. The intrinsic glucose affinity to the activator-bound enzyme demonstrated a 700-fold increase relative to the apo-enzyme. This is consistent with a reduction in apparent glucose KD and the steady-state parameter K0.5 as a result of enzyme equilibrium shifting to the activator-bound form. The binding of small molecules to GK was dependent on glucose, consistent with the structural evidence for an allosteric binding site which is present in the glucose-induced, active-enzyme form of GK, and absent in the inactive apo-enzyme. A mechanistic model that brings together the kinetic and structural data is proposed which allows qualitative and quantitative analysis of the glucose-dependent GK regulation by small molecules. The regulation of GK activation by glucose may have an important implication for the discovery and design of GK activators as potential antidiabetic agents.

Support was provided by the Pfizer La Jolla Postdoctoral Program.





This Article
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