A transient kinetic study on the reactivity of recombinant unprocessed monomeric myeloperoxidase

Spectral and kinetic features of the redox intermediates of human recombina nt unprocessed monomeric myeloperoxidase (recMPO), purified from an enginee red Chinese hamster ovary cell line, were studied by the multi-mixing stopp ed-flow technique. Both the ferric protein and compounds I and II showed es sentially the same kinetic behavior as the mature dimeric protein (MPO) iso lated from polymorphonuclear leukocytes. Firstly, hydrogen peroxide mediate d both oxidation of ferric recMPO to compound I (1.9 x 10(7) M-1 s(-1), pH 7 and 15 degreesC) and reduction of compound I to compound II (3.0 x 10(4) M-1 s(-1), pH 7 and 15 degreesC). With chloride, bromide, iodide and thiocy anate compound I was reduced back to the ferric enzyme (3.6 x 10(4) M-1 s(- 1), 1.4 x 10(6) M-1 s(-1), 1.4 x 10(7) M-1 s(-1) and 1.4 x 10(7) M-1 s(-1), respectively), whereas the endogenous one-electron donor ascorbate mediate d transformation of compound I to compound II (2.3 x 10(5) M-1 s(-1)) and o f compound II back to the resting enzyme (5.0 x 10(3) M-1 s(-1)). Comparing the data of this study with those known from the mature enzyme strongly su ggests that the processing of the precursor enzyme (recMPO) into the mature form occurs without structural changes at the active site and that the sub units in the mature dimeric enzyme work independently. (C) 2001 Federation of European Biochemical Societies. Published by Elsevier Science BY. All ri ghts reserved.