COMPARISON OF VIBRATIONAL FREQUENCIES OF CRITICAL BONDS IN GROUND-STATE COMPLEXES AND IN A VANADATE-BASED TRANSITION-STATE ANALOG COMPLEX OF MUSCLE PHOSPHOGLUCOMUTASE - MECHANISTIC IMPLICATIONS

The Symmetric stretching frequency of the P-O bonds of the enzymic phosphate group in muscle phosphoglucomutase was measured via O-16/O-18 Raman difference spectroscopy. This frequency, and its shift on isotopic substitution, is characteristic of a dianionic phosphate ester. The P-O stretching frequency is not detectably altered by the binding of the metal ion activators Mg2+, Zn2+, or Cd2+ nor by the subsequent binding of glucose phosphate. Hence, a binding-induced distortion/polarization of the enzymic phosphate group in the ground state, or enzyme-substrate complex, cannot serve as a rationale for the large value of k(cat) in the phosphoglucomutase reaction. By contrast, the stretching frequency of the V-O bonds within a vanadate group bound at the same site in the transition-state analog complex involving glucose 1-phosphate 6-vanadate is much lower than for a normal dianionic vanadate. This low V-O stretching frequency is best rationalized in terms of the extensive polarization of all three nonbridging oxygens of the vanadate ester dianion plus the formation of a weak, fifth bond to the vanadium atom. This distortion/polarization of the VO32- group depends on the metal ion activator, since it is largely abolished, and the involvement of the fifth ligand eliminated, by substitution of Li+ for Mg2+ at the metal activation site. To the extent that the vanadate-inhibitor complex mimics the transition state for the normal phosphoglucomutase reaction, as has been suggested [Ray, W. J., Jr., & Puvathingal, J. M. (1990) Biochemistry 29, 2790], the normal PO3- transfer is best described as a process with S(N)2-like or associative character and thus is quite different from the process by which model phosphate ester dianions normally react in aqueous solution.