The potential role of nitric oxide in substrate switching in eosinophil peroxidase.
Eosinophil recruitment and enhanced nitric oxide (NO) production are characteristic features of asthma and other airway diseases. Eosinophil peroxidase (EPO), a highly cationic hemoprotein secreted by activation of eosinophils, is believed to play a central role in host defense against invading pathogens. The enzyme uses hydrogen peroxide (H2O2) and bromide (Br-), a preferred cosubstrate of EPO, to generate the cytotoxic oxidant hypobromous acid. The aim of this work was to determine whether NO can compete with plasma levels of Br- and steer the enzyme reaction from a 2e- oxidation to a 1e- oxidation pathway. Rapid kinetic measurements were utilized to measure the rate of EPO compounds I and II formation, duration, and decay at 412 and 432 nm, respectively, at 10 degrees C. An EPO-Fe(III) solution supplemented with increasing Br- concentrations was rapidly mixed with fixed amounts of H2O2 in the absence and in the presence of increasing NO concentrations. In the absence of NO, EPO-Fe(III) primarily converted to compound I and, upon H2O2 exhaustion, it decayed rapidly to the ferric form. NO caused a significant increase in the accumulation of EPO compound II, along with a proportional increase in its rate of formation and duration as determined by the time elapsed during catalysis. The time courses for these events have been incorporated into a comprehensive kinetic model. Computer simulations carried out supported the involvement of a conformational intermediate in the EPO compound II complex decay. Collectively, our results demonstrated that NO displays the potential capacity to promote substrate switching by modulating substrate selectivity of EPO.
Galijasevic
Semira
S
Department of Obstetrics and Gynecology, The C.S. Mott Center for Human Growth and Development, Children's Hospital of Michigan, Wayne State University School of Medicine, Detroit, Michigan 48201, USA
Proteasa
Gheorghe
G
Abdulhamid
Ibrahim
I
Abu-Soud
Husam M
HM
D000818:Animals; D001965:Bromides / Q000378:metabolism; D047209:Eosinophil Peroxidase / Q000097:blood / Q000737:chemistry* / Q000378:metabolism*; D004804:Eosinophils / Q000201:enzymology; D066298:In Vitro Techniques; D007700:Kinetics; D008958:Models, Molecular; D009569:Nitric Oxide / Q000378:metabolism; D010084:Oxidation-Reduction; D011487:Protein Conformation; D013053:Spectrophotometry; D013379:Substrate Specificity; D013552:Swine
D001965:Bromides; D009569:Nitric Oxide; D047209:Eosinophil Peroxidase