Myeloperoxidase interaction with peroxynitrite: chloride deficiency and heme depletion

Abstract

Myeloperoxidase (MPO) is a hemoprotein, involved in the leukocyte mediated defense mechanism, and uses hydrogen peroxide (H2O2) and chloride (Cl-) to produce hypochlorous acid. In human saliva and hypochloremic alkalosis syndrome occurring in breast fed infants, the MPO-H2O2 system functions in lower Cl- concentration (10-70 mM) compared to plasma levels (100 mM) as part of the antibacterial defense system. The impact of low Cl- concentration and exposure to high peroxynitrite (ONOO-) synthesized from cigarette smoke or oxidative stress on MPO function is still unexplored. Rapid mixing of ONOO- and MPO caused immediate formation of a transient intermediate MPO Compound II which then decayed to MPO-Fe (III). Double mixing of MPO with ONOO- followed by H2O2 caused immediate formation of Compound II followed by MPO heme depletion, a process that occured independent of ONOO- concentration. Peroxynitrite/H2O2-mediated MPO heme depletion was confirmed by HPLC analysis and in-gel heme staining showing 60-70% less heme content compared to the control. A non-reducing denaturing SDS PAGE showed no fragmentation or degradation of protein. Myeloperoxidase heme loss was completely prevented by pre-incubation of MPO with saturated amounts of Cl-. Chloride binding to the active site of MPO constrains ONOO- binding by filling the space directly above the heme moiety or by causing a protein conformational change that constricts the distal heme pocket, thus preventing ONOO- from binding to MPO heme iron. Peroxynitrite interaction with MPO may serve as a novel mechanism for modulating MPO catalytic activity, influencing the regulation of local inflammatory and infectious events in vivo.

Impact of hydrogen peroxide-driven Fenton reaction on mouse oocyte quality

“…In biological system, the source of free iron that is capable of catalyzing free radical reactions is still not known, but it thought to be generated from iron storage proteins, mitochondria and heme proteins such as MPO, hemoglobin and myoglobin [17–19, 21, 22, 36–38]. Recent studies have shown that MPO-HOCl system not only play an important role in generating •OH [10] through the formation of ferryl ion intermediate, but it also serves as a source of free iron through its ability to mediate hemoprotein heme destruction [20–22].…”

“…Previously, we have demonstrated that MPO can serve as a source of free iron under conditions where both peroxynitrite and H 2 O 2 are elevated [19]. Recently, we have shown that HOCl, the final product of MPO, could mediate-hemoprotein heme destruction and release free iron [20–22].…”

The reaction of HOCl and cyanocobalamin: Corrin destruction and the liberation of cyanogen chloride

“…HOCl-mediated Cobl destruction may also disturb the conversion of methylmalonic acid to succinyl-CoA [49], an input to the citric acid cycle [50, 51]. Previously, we have shown that MPO may serve as a source of free iron under oxidative stress when both NO and O 2 •− are elevated [52]. It has also been shown that iron accumulates in atherosclerotic lesions in a catalytically active form.…”

Early Detection Of Breast Cancer using Post-Translationally Modified Biomarkers

“…Because tyrosines that are post-translationally modified by nitrating oxidants, brominating oxidants, and chloramination oxidants produced from macrophages, neutrophils or eosinophils to form nitrotyrosine, bromotyrosine and chlorotyrosine respectively, have relevance to breast cancers (17)(18)(19)(20)(21)(22)(23). For example, hypochlorous acid/hypochlorite adducts have been reported for breast cancer (24)(25)(26)(27). Our studies suggest that circulating PTM levels can be used as a biomarker for endothelial cell dysfunction, which is of concern in several human diseases.…”