Ferric nitrosylated myoglobin catalyzes peroxynitrite scavenging.

Myoglobin (Mb), generally taken as the molecular model of monomeric globular heme-proteins, is devoted: (i) to act as an intracellular oxygen reservoir, (ii) to transport oxygen from the sarcolemma to the mitochondria of vertebrate heart and red muscle cells, and (iii) to act as a scavenger of nitrogen and oxygen reactive species protecting mitochondrial respiration. Here, the first evidence of (.)NO inhibition of ferric Mb- (Mb(III)) mediated detoxification of peroxynitrite is reported, at pH 7.2 and 20.0 degrees C. (.)NO binds to Mb(III) with a simple equilibrium; the value of the second-order rate constant for Mb(III) nitrosylation (i.e., (.NO)kon) is (6.8 +/- 0.7) x 10(4) M(-1) s(-1) and the value of the first-order rate constant for Mb(III)-NO denitrosylation (i.e., (.NO)koff) is 3.1 +/- 0.3 s(-1). The calculated value of the dissociation equilibrium constant for Mb(III)-NO complex formation (i.e., (.NO)koff/(.NO)kon = (4.6 +/- 0.7) x 10(-5) M) is virtually the same as that directly measured (i.e., (.NO)K = (3.8 +/- 0.5) x 10(-5) M). In the absence of (.)NO, Mb(III) catalyzes the conversion of peroxynitrite to NO3(-), the value of the second-order rate constant (i.e., (P)kon) being (1.9 +/- 0.2) x 10(4) M(-1) s(-1). However, in the presence of (.)NO, Mb(III)-mediated detoxification of peroxynitrite is only partially inhibited, underlying the possibility that also Mb(III)-NO is able to catalyze the peroxynitrite isomerization, though with a reduced rate ((P)kon* = (2.8 +/- 0.3) x 10(3) M(-1) s(-1)). These data expand the multiple roles of (.)NO in modulating heme-protein actions, envisaging a delicate balancing between peroxynitrite and (.)NO, which is modulated through the relative amount of Mb(III) and Mb(III)-NO.