EPR determination of the Co(II)-free radical magnetic geometry of the "doublet" species arising in a coenzyme B-12-enzyme reaction.

The physical significance of the observed structure of the EPR signal, commonly known as the "doublet" spectrum, is that it contains information not only about the exchange coupling but also about the geometry of the magnetic dipole-dipole spin-spin coupling. We can show this because we have developed a general method of analysis applicable to this type of system and because we demand a quantitative fit of theory to experiment at two microwave frequencies. We have chosen the "doublet" free radical signal, which arises in the ribonucleotide reductase-5'-deoxyadenosylcobalamin system (from Lactobacillus leichmannii, see Hamilton et al., Biochemistry 11, 4696--4705 (1972)), for study, for the particular reason that the 35 GHz "doublet" spectrum has three components (in this case) rather than two, which provides an important test of the recently proposed model of isotropic exchange coupling by Schepler et al. ((1975) Biochim. Biophys. Acta 397, 510--518). We find that a quantitative fit to the EPR "doublet" lineshape can be obtained with a model of isotropic exchange, and a "point" magnetic dipole-dipole interaction acting over a distance of 9.9 A with the radical located approx. 34 degrees off the principal gzz axis and less than 1 degree off the principal gxx axis of the Co(II) in the corrin ring. Quantitative fits of the doublet portion of the observed lineshape at both 9 and 35 GHz were achieved with this model, assuming an axially symmetric free radical signal and a Gaussian spin-packet lineshape with isotropic linewidth.