GLUCOSE ANALOG INHIBITORS OF GLYCOGEN-PHOSPHORYLASE - FROM CRYSTALLOGRAPHIC ANALYSIS TO DRUG PREDICTION USING GRID FORCE-FIELD AND GOLPE VARIABLE SELECTION

Several inhibitors of the large regulatory enzyme glycogen phosphoryla se (GP) have been studied in crystallographic and kinetic experiments. GP catalyses the first step in the phosphorylysis of glycogen to gluc ose-1-phosphate, which is utilized via glycolysis to provide energy to sustain muscle contraction and in the liver is converted to glucose. alpha-D-Glucose is a weak inhibitor of glycogen phosphorylase form b ( GPb, K-i = 1.7 mM) and acts as a physiological regulator of hepatic gl ycogen metabolism. Glucose binds to phosphorylase at the catalytic sit e and results in a conformational change that stabilizes the inactive T state of the enzyme, promoting the action of protein phosphatase 1 a nd stimulating glycogen synthase. It has been suggested that in the li ver, glucose analogues with greater affinity for glycogen phosphorylas e may result in a more effective regulatory agent. Several N-acetyl gl ucopyranosylamine derivatives have been synthesized and tested in a se ries of crystallographic and kinetic binding studies with GPb. The str uctural results of the bound enzyme-ligand complexes have been analyse d together with the resulting affinities in an effort to understand an d exploit the molecular interactions that might give rise to a better inhibitor. Comparison of the N-methylacetyl glucopyranosylamine (N-met hylamide, K-i = 0.032 mM) with the analogous beta-methylamide derivati ve (C-methylamide, K-i = 0.16 mM) illustrate the importance of forming good hydrogen bonds and obtaining complementarity of van der Waals in teractions. These studies also have shown that the binding modes can b e unpredictable but may be rationalized with the benefit of structural data and that a buried and mixed polar/nonpolar catalytic site poses problems for the systematic addition of functional groups. Together wi th previous studies of glucose analogue inhibitors of GPb, this work f orms the basis of a training set suitable for three-dimensional quanti tative structure-activity relationship studies. The molecules in the t raining set are void of problems and potential errors arising from the alignment and bound conformations of each of the ligands since the co ordinates were those determined experimentally from the X-ray crystall ographic refined ligand-enzyme complexes. The computational procedure described in this work involves the use of the program GRID to describ e the molecular structures and the progam GOLPE to obtain the partial least squares regression model with the highest prediction ability. Th e GRID/GOLPE procedure performed using 51 glucose analogue inhibitors of GPb has good overall predictivity [standard deviation of error pred ictions (SDEP) = 0.98 and Q(2) = 0.76] and has shown good agreement wi th the crystallographic and kinetic results by reliably selecting regi ons that are known to affect the binding affinity.