Quantitation of movement of the phosphoryl group during catalytic transfer in the arginine kinase reaction: ³¹P relaxation measurements on enzyme-bound equilibrium mixtures^**

Abstract

³¹P nuclear spin relaxation measurements have been made on enzyme-bound equilibrium mixtures of lobster-muscle arginine kinase in the presence of substituent activating paramagnetic cation Co(II) (in place of Mg(II)), i.e., on samples in which the reaction, E•CoATP•arginine ⇌ E•CoADP•P-arginine, is in progress. The results have been analyzed on the basis of a previously published theory (Nageswara Rao, B.D. (1995) J. Magn. Reson., B108, 289–293) to determine the structural changes in the reaction complex accompanying phosphoryl transfer. The analysis enables the determination of the change in the Co(II)-³¹P (γ-P(ATP)) vector as the transferable phosphoryl group moves over and attaches to arginine to form P-arginine. It is shown that the Co(II)-³¹P distance of ∼3.0 Å, representing direct coordination of Co(II) to γ-P(ATP), changes to ∼4.0 Å when P-arginine is formed in the enzyme-bound reaction complex. This elongation of the Co(II)-³¹P vector implies an excursion of at least 1.0 Å for the itinerant phosphoryl group on the surface of the enzyme.