THE COENZYME B-12 DERIVATIVE N1-METHYL-5'-DEOXYADENOSYLCOBALAMIN - SYNTHESIS, CHARACTERIZATION, STABILITY TOWARDS DIMROTH REARRANGEMENT, AND CO-C THERMOLYSIS PRODUCT AND KINETIC-STUDIES

The mechanism of Co-C heterolysis of adenosylcobalamin (AdoCbl) is stu died, specifically the effect of a positive charge produced by methyla tion (CH:, a ''sticky proton'' analog of H+ addition) to the most basi c adenine N1 nitrogen site in AdoCbl. This is accomplished by the synt hesis, characterization and Co-C thermolysis in the presence of the ra dical trap TEMPO of the N1 methylated AdoCbl derivative, [N1-methyl-5' -deoxyadenosycobalamin](+), 5. A variety of needed additional synthese s and other control experiments are also reported, including effective purification methods using ultrafiltration membranes, procedures that may prove to be the more generally useful part of the synthetic work reported. The thermolysis results with 5 are unequivocal in showing th at a positive charge at the N1 nitrogen in the adenosyl group of AdoCb l has no effect within +/-8% on the products or rates of Co-C thermal cleavage. The data are most consistent with a recently suggested, new mechanism for AdoCbl Co-C heterolysis at less acidic pH values, one no t involving the protonation of the beta-ribose oxygen that is known to occur under strongly acidic conditions. In addition, the present stud y illustrates some of the positive features, and also some of the pitf alls, in the approach where CH3+ is used as a ''sticky proton'' analog in mechanistic studies. (C) 1998 Elsevier Science Inc. All rights res erved.