Mycobacterium leprae truncated hemoglobin O (trHbO) protects from nitrosative stress and sustains mycobacterial respiration. Here, kinetics of M. leprae trHbO(II)-NO denitrosylation and of O-2-mediated oxidation of M. leprae trHbO(II)-NO are reported. Values of the first-order rate constant for (NO)-N-center dot dissociation from M. leprae trHbO(II)-NO (k(off)) and of the first-order rate constant for O-2-mediated oxidation of M. leprae trHbO(II)-NO (h) are 1.3 x 10(-4) s(-1) and 1.2 x 10(-4) s(-1), respectively. The coincidence of values of k(off) and h suggests that O-2-mediated oxidation of M. leprae trHbO(II)-NO occurs with a reaction mechanism in which (NO)-N-center dot, that is initially bound to heme(II), is displaced by O-2 but may stay trapped in a protein cavity(ies) close to heme(II). Next, M. leprae trHbO(II)-O-2 reacts with (NO)-N-center dot giving the transient Fe(III)-OONO species preceding the formation of the final product M. leprae trHbO(III). (center dot)NOheme(II)-NO represents the rate limiting step for O-2-mediated oxidation of M. leprae trHbO(II)-NO. (c) 2007 Elsevier Inc. All rights reserved.