OXIDATION OF THREONYLMETHIONINE BY PEROXYNITRITE - QUANTIFICATION OF THE ONE-ELECTRON TRANSFER PATHWAY BY COMPARISON TO ONE-ELECTRON PHOTOOXIDATION

Peroxynitrite can modify methionine by one- and two-electron oxidation pathways. Here, we have quantified the extent of one-electron oxidati on of threonylmethionine (Thr-Met) by peroxynitrite using a characteri stic reaction according to which Thr-Met sulfur radical cations decomp ose via fragmentation of the Thr side chain, yielding acetaldehyde. Th e efficiencies, f(acet,photo), for the formation of acetaldehyde from Thr-Met sulfur radical cations were obtained by means of one-electron photooxidation using triplet 4-carboxybenzophenone. Exact quantum yiel ds for the formation of Thr-Met sulfur radical cations by triplet 4-ca rboxybenzophenone were obtained by laser flash photolysis and time-res olved UV spectroscopy. Acetaldehyde yields were measured for the react ion of peroxynitrite with Thr-Met, and division of these acetaldehyde yields by f(acet,photo) yielded the extents to which peroxynitrite rea cted with Thr-Met via the one-electron transfer pathway. There was lit tle one-electron oxidation of Thr-Met by peroxynitrite at pH 7.4, i.e. , 1.5%, 1.8%, and 5.3% based on the total chemical conversion of Thr-M et for Thr-Met concentrations of 1 x 10(-3), 5 x 10(-4), and 1.75 x 10 (-4) M, respectively. In all cases the major reaction product was the two-electron oxidation product threonylmethionine sulfoxide. However, at pH 6.0, one-electron oxidation of Thr-Met showed a significantly hi gher efficiency of 14% for [Thr-Met] = 1.75 x 10(-4) M. Under all expe rimental conditions the extent of one-electron oxidation increased wit h decreasing peptide concentration in agreement with a recently establ ished mechanism according to which the one-electron oxidation of Met b y peroxynitrite requires a unimolecular transformation of peroxynitrou s acid to an excited species which is the ultimate one-electron oxidan t.