19F NMR investigations of the catalytic mechanism of phosphoglucomutase using fluorinated substrates and inhibitors.

The complexes of phosphoglucomutase with a number of fluorinated substrate analogues have been investigated by 19F NMR and the effects of the binding of Li+ and Cd2+ to these complexes determined. Very large downfield chemical shift changes (-14 to -19 ppm) accompanied binding of the inhibitors 6-deoxy-6-fluoro-alpha-D-glucopyranosyl phosphate and alpha-glucosyl fluoride 6-phosphate to the phosphoenzyme. Smaller shift changes were observed for ligands substituted with fluorine at other positions. Addition of Li+ to enzyme/fluorinated ligand complexes caused a 10(2)- to 10(3)-fold decrease in ligand dissociation constants as witnessed by the change from intermediate to slow-exchange conditions in the NMR spectra. Measurement of the 19F NMR spectra of complexes of the Li(+)-enzyme with each of the fluoroglucose 1-phosphates and 6-phosphates has provided some insight into the environment of each of these fluorines (thus also parent hydroxyls) in each of the complexes. Results obtained argue strongly against a single sugar binding mode for the glucose 1- and 6-phosphates. Two enzyme-bound species were detected in the 19F NMR spectra of the complexes formed by reaction of the Cd(2+)-phosphoenzyme complex with the 2- and 3-fluoroglucose phosphates. These are tentatively assigned as the fluoroglucose 1,6-bisphosphate species bound in two different modes to the dephosphoenzyme. Only one bound species was observed in the case of the 4-fluoroglucose phosphates.(ABSTRACT TRUNCATED AT 250 WORDS)