ELECTRON-PARAMAGNETIC RESONANCE STUDIES OF A RAS P21-MN(II) GDP COMPLEX IN SOLUTION

The number of water molecules bound to Mn2+ in the complex with a variant of Ha ras p21 and GDP has been determined by electron paramagnetic resonance (EPR) measurements in O-17-enriched water. A resolution enhancement method has been used to improve quantitation of the spectral data. These spectroscopic measurements show that Mn2+ has four water ligands in this complex, a result in agreement with the conclusions of a previous paper [Smithers, G. W., Poe, M., Latwesen, D. G., & Reed, G. H. (1990) Arch. Biochem. Biophys. 280, 416-420]. The resolution enhancement method has also been applied in a measurement of the O-17-Mn2+ superhyperfine coupling constant of O-17 in the beta-phosphate of the GDP in the ras p21 complex. The intrinsically narrow EPR signals of Mn2+ in the complex with ras p21 and GDP in (H2O)-H-2 respond to resolution enhancement such that the superhyperfine splitting from the O-17 nuclear spin (I = 5/2) becomes visible in the EPR signals. An O-17-Mn2+ superhyperfine coupling constant is obtained from simulation of the resolution-enhanced EPR spectrum.