TIME-RESOLVED SPECTROSCOPIC STUDIES OF B-12 COENZYMES - THE PHOTOLYSIS AND GEMINATE RECOMBINATION OF ADENOSYLCOBALAMIN

Femtosecond transient absorption spectroscopy has been used to investi gate the photolysis of coenzyme B-12 (5'-deoxyadenoSylcobalamin). Tran sient kinetic measurements obtained at wavelengths between 400 and 633 nm were analyzed globally to obtain time constants. Transient spectra l data obtained for time delays between 5 ps and 9 ns were analyzed by matrix decomposition to identify distinct spectral components present in the data. Photoexcitation results in homolysis of the carbon-cobal t bond forming a singlet radical pair on a picosecond time scale. The subsequent spectral changes probe conformational relaxation and gemina te recombination. Analysis of the spectral data suggests that 76 +/- 4 % of the geminate radical pairs recombine, resulting in a quantum yiel d of 0.24 +/- 0.04 for the formation of solvent separated radicals, in good agreement with literature values of 0.20 +/- 0.03 and 0.23 +/- 0 .04 [Chen, E.; Chance, M. R. Biochemistry 1993, 32, 1480-1487]. The ge minate recombination of the adenosyl radical with cob(II)alamin occurs biphasically with exponential time constants of 150 +/- 20 ps and 0.5 +/- 0.2 ns. The effective recombination rate from a single-exponentia l fit to the data is (0.250 ns)(-1) = 4 ns(-1).