Time-Resolved Spectroscopic Studies of B₁₂ Coenzymes:  The Photolysis of Methylcobalamin Is Wavelength Dependent

Abstract

Femtosecond to nanosecond transient absorption spectroscopy has been used to investigate the primary photochemistry of the B 12 coenzymes, methylcobalamin and 5′-deoxyadenosylcobalamin. Photolysis at excitation wavelengths in the near UV (400 nm) and visible (520-530 nm) are compared. Measurements were performed with femtosecond time resolution covering time delays of up to 9 ns. The photochemistry of methylcobalamin is found to depend strongly on excitation wavelength, while the photochemistry of adenosylcobalamin is essentially wavelength independent over the range studied. Excitation of methylcobalamin at 400 nm results in a partitioning between prompt bond homolysis and formation of a metastable cob(III)-alamin photoproduct as reported earlier [Walker, L. A., II; Jarrett, J. T.; Anderson, N. A.; Pullen, S. H.; Matthews, R. G.; Sension, R. J. J. Am. Chem. Soc. 1998, 120, 3597-3603]. Excitation of methylcobalamin at 520 nm in the visible R -band results only in formation of the metastable cob(III)alamin photoproduct. No prompt bond homolysis is observed. The metastable photoproduct partitions between formation of cob(II)-alamin (14 ( 5%) and recovery of the methylcobalamin starting material (86 ( 5%) on a 1.0 ( 0.1 ns time scale. The quantum yield for bond homolysis in methylcobalamin is determined by the wavelength-dependent partitioning between prompt homolysis and formation of the metastable photoproduct and by the partitioning of the metastable photoproduct between bond homolysis and ground-state recovery. In contrast, excitation of adenosylcobalamin at both 400 and 520 nm results in the development of a difference spectrum characteristic of the formation of cob(II)alamin on a picosecond time scale. The quantum yield for bond homolysis in this case is determined primarily by the competition between geminate recombination and diffusion to form solventseparated radical pairs. The caging fraction for adenosylcobalamin in aqueous solution at room temperature is 0.71 ( 0.05.

The Nature of the Co-C Bond Cleavage Processes in Methylcob(II)Alamin and Adenosylcob(III)Alamin

“…As it is depicted in Figure 3: (a) the methylcobalamin-dependent methionine synthase generally includes as a primary turnover the cycling between Co(III) methylcobalamin and cob(I)alamin, which allows the heterolytic Co-C cleavage process, while in the adenosyl-cobalamin-dependent mutases the Co-C bond of adenosylcobalamin is cleaved homolytically, giving rise to a fi ve-coordinate cob(II)alamin and an adenosyl radical; (b) the association of the substrate and adenosyl-ligand during the Co-C cleavage process in adenosylcobalamin dependent mutases has been proved by various experimental data including X-ray diffraction results [35][36][37], while such effect is totally absent in studies regarding methylcobalamindependent mutases. The Co-C cleavage and electron transfer processes have been studied by different methods such as electrochemistry [38][39][40], thermo- [41] or photochemistry [42][43][44][45][46] of methylcobalamin Co(III) by a signifi cant number of researchers. As a result, many triggering factors of Co-C bond breaking have been considered and various mechanistic mechanisms of the Co-C cleavage process have been proposed.…”

Theoretical investigations of photophysics and spectroscopic properties in B12 cofactors.

“…56,58,60,65 To The dynamics of the photoinduced homolysis of B 12 cofactors were investigated on a time scale ranging from femtoseconds to nanoseconds, allowing for the detailed mechanism of Co-C bond photolysis to be obtained. In the case of CNCbl, it was found that the metastable S 1 state has a ligand-to-metal charge transfer (LMCT)…”

“…However, the excitation at 520 nm was followed only by formation of a long-lived S 1 state. 56,58,65 In contrast, the photolysis of AdoCbl was found to be wavelength independent resulting in a creation of a geminate radical pair within 100-130 ps at room temperature. 57,59,62 The same studies for ethyland n-propylcobalamin indicated the formation of radical pair when samples were excited at both 400 and 520 nm but the rapid dissociation contribution was mainly observed following the higher energy excitation.…”

“…The most recent results originate from the TAS studies by Sension et al 56,58,60,65 and the spectroscopic studies by…”

“…Whereas the metastable S 1 state in CNCbl was assigned as corresponding to ligand-to-metal charge transfer (LMCT) transition, 64 the S 1 in MeCbl was described as metal-to-ligand charge transfer (MLCT). 56,58,60 On the other hand, the S 1 for coenzyme B 12 presented different character (MLCT or LMCT) depending on the environment. 65 In the light of the above findings, the photochemistry of cobalamins was further explored with the aim of theoretical studies.…”