STRUCTURE OF THE NOVEL HEME ADDUCT FORMED DURING THE REACTION OF HUMAN HEMOGLOBIN WITH BRCCL3 IN RED-CELL LYSATES

It was previously shown that the reductive debromination of BrCCl3 to trichloromethyl radical by human hemoglobin leads to formation of diss ociable altered heme products, two of which are identical to those for med from myoglobin and one which is novel. In this study, we have eluc idated the structure of this novel adduct with the use of mass spectro metry, as well as H-1 and C-13 NMR as a substitution product of a -C(C l)=CCl2 moiety for a beta-hydrogen atom on the prosthetic heme's ring I vinyl group. From studies with the use of C-13-enriched BrCCl3, it w as determined that the added carbon atoms were derived from 2 eq of Br CCl3. A mechanism that involves multiple reductive events and a radica l cation heme intermediate is proposed. Consistent with this mechanism , cellular reductants were found to selectively enhance the amount of this novel dissociable heme adduct. These studies reveal fine differen ces between myoglobin and hemoglobin in the accessibility of reactive intermediates to the ring I vinyl group, as well as the potential impo rtance of cellular reductants on the course of heme alteration.