Myeloperoxidase-dependent fluorescein chlorination by stimulated neutrophils.

Hurst, James K.; Albrich, J M; Green, T R; Rosen, Henry; Klebanoff, Seymour J.

doi:10.1016/s0021-9258(17)42918-8

Cited by 94 publications

(45 citation statements)

References 31 publications

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“…Amino acid residues such as tyrosine, tryptophan, lysine, methionine, and cysteine are common targets for oxidative modifications, including halogenation, hydroxylation, nitration, nitrosylation, carbamylation, oxidative cross-links, and elevated states of sulfur oxidation . In particular, post-translational modifications (PTM) of tyrosine residues have emerged as biomarkers for oxidative damage that convey chemical information about the oxidative process involved in their generation and have been observed in more than 50 human pathologies. − In neutrophils and eosinophils, myeloperoxidase (MPO) and eosinophil peroxidase, respectively, mediate the formation predominantly of hypochlorous and hypobromous acids. − In macrophages, activation of NADPH oxidase and inducible nitric oxide synthase (iNOS) results in the concurrent formation of superoxide and nitric oxide (NO), which together form peroxynitrite (ONOO – ). , These reactive oxygen species (ROS) and reactive nitrogen species (RNS) are potent tyrosine oxidants, forming chloroTyr, bromoTyr, and nitroTyr under oxidative stress conditions. ,,− Site-specific tyrosine nitration and chlorination have been shown to affect protein–protein interactions with detrimental downstream effects on cellular function and have been established as important biomarkers of oxidative disease states. − However, the specific effects of tyrosine halogenation on protein function have not been well documented. It is proposed that these oxidative stress-induced post-translational modifications (ox-PTMs) alter protein function and protein–protein interactions …”

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confidence: 99%

Efficient Site-Specific Prokaryotic and Eukaryotic Incorporation of Halotyrosine Amino Acids into Proteins

Jang

Cooley

et al. 2020

ACS Chem. Biol.

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Post-translational modifications (PTMs) of protein tyrosine (Tyr) residues can serve as a molecular fingerprint of exposure to distinct oxidative pathways and are observed in abnormally high abundance in the majority of human inflammatory pathologies. Reactive oxidants generated during inflammation include hypohalous acids and nitric oxide-derived oxidants, which oxidatively modify protein Tyr residues via halogenation and nitration, respectively, forming 3-chloroTyr, 3-bromoTyr, and 3-nitroTyr. Traditional methods for generating oxidized or halogenated proteins involve nonspecific chemical reactions that result in complex protein mixtures, making it difficult to ascribe observed functional changes to a site-specific PTM or to generate antibodies sensitive to site-specific oxidative PTMs. To overcome these challenges, we generated a system to efficiently and site-specifically incorporate chloroTyr, bromoTyr, and iodoTyr, and to a lesser extent nitroTyr, into proteins in both bacterial and eukaryotic expression systems, relying on a novel amber stop codon-suppressing mutant synthetase (haloTyrRS)/tRNA pair derived from the Methanosarcina barkeri pyrrolysine synthetase system. We used this system to study the effects of oxidation on HDL-associated protein paraoxonase 1 (PON1), an enzyme with important antiatherosclerosis and antioxidant functions. PON1 forms a ternary complex with HDL and myeloperoxidase (MPO) in vivo. MPO oxidizes PON1 at tyrosine 71 (Tyr71), resulting in a loss of PON1 enzymatic function, but the extent to which chlorination or nitration of Tyr71 contributes to this loss of activity is unclear. To better understand this biological process and to demonstrate the utility of our GCE system, we generated PON1 site-specifically modified at Tyr71 with chloroTyr and nitroTyr in Escherichia coli and mammalian cells. We demonstrate that either chlorination or nitration of Tyr71 significantly reduces PON1 enzymatic activity. This tool for site-specific incorporation of halotyrosine will be critical to understanding how exposure of proteins to hypohalous acids at sites of inflammation alters protein function and cellular physiology. In addition, it will serve as a powerful tool for generating antibodies that can recognize site-specific oxidative PTMs.

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confidence: 99%

Efficient Site-Specific Prokaryotic and Eukaryotic Incorporation of Halotyrosine Amino Acids into Proteins

Jang

Cooley

et al. 2020

ACS Chem. Biol.

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“…Exclusively in neutrophils, MPO uses H 2 O 2 , Cl 2 , and H + to produce HOCl [44]. HOCl is the most potent oxidant produced by neutrophils, because it reacts with a broad spectrum of macromolecules in cells [45][46][47][48][49][50][51]. However, the question of how phagosomes acquire chloride for such an essential biosynthesis remains unresolved.…”

Section: Discussionmentioning

confidence: 99%

CFTR targeting during activation of human neutrophils

Valentine

Wang

2016

Journal of Leukocyte Biology

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Cystic fibrosis transmembrane conductance regulator (CFTR), a cAMP-activated chloride channel, plays critical roles in phagocytic host defense. However, how activated neutrophils regulate CFTR channel distribution subcellularly is not well defined. To investigate, we tested multiple Abs against different CFTR domains, to examine CFTR expression in human peripheral blood neutrophils by flow cytometry. The data confirmed that resting neutrophils had pronounced CFTR expression. Activation of neutrophils with soluble or particulate agonists did not significantly increase CFTR expression level, but induced CFTR redistribution to cell surface. Such CFTR mobilization correlated with cell-surface recruitment of formyl-peptide receptor during secretory vesicle exocytosis. Intriguingly, neutrophils from patients with ΔF508-CF, despite expression of the mutant CFTR, showed little cell-surface mobilization upon stimulation. Although normal neutrophils effectively targeted CFTR to their phagosomes, ΔF508-CF neutrophils had impairment in that process, resulting in deficient hypochlorous acid production. Taken together, activated neutrophils regulate CFTR distribution by targeting this chloride channel to the subcellular sites of activation, and ΔF508-CF neutrophils fail to achieve such targeting, thus undermining their host defense function.

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“…A pioneering study reported that neutrophil phagosomes alkalinized a phenotype that was attributed to the consumption of proton by the dismutation reaction of superoxide to H 2 O 2 [33]. This study, however, was subsequently criticized, as it failed to consider the MPO-dependent chlorination of the pH sensor fluorescein within neutrophil phagosomes [37]. A more recent study indicated that neutrophil phagosomes remain neutral, as V-ATPases do not accumulate efficiently on phagosomes when the oxidase is active [24].…”

Section: Discussionmentioning

confidence: 99%

Hv1 proton channels differentially regulate the pH of neutrophil and macrophage phagosomes by sustaining the production of phagosomal ROS that inhibit the delivery of vacuolar ATPases

Chemaly

Nunes

Jimaja

et al. 2014

Journal of Leukocyte Biology

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Production of ROS and maintenance of an appropriate pH within the lumen of neutrophil and macrophage phagosomes are important for an effective immune response. Hv1 proton channels sustain ROS production at the plasma membrane, but their role in phagosomes is not known. Here, we tested whether Hv1 channels regulate the pH and sustain phagosomal ROS production in neutrophils and macrophages. The presence of Hv1 channels on phagosomes of human neutrophils and mouse macrophages was confirmed by Western blot and immunostaining. Phagosomal ROS production, measured with OxyBurst-coupled targets, was reduced in neutrophils and macrophages isolated from Hv1-deficient mice. Ratiometric imaging of FITC-coupled targets showed that phagosomes acidified more slowly in Hv1-deficient macrophages and transiently alkalinized when the V-ATPase was inhibited. In WT neutrophils, 97% of phagosomes remained neutral 30 min after particle ingestion, whereas 37% of Hv1-deficient phagosomes were alkaline (pH>8.3) and 14% acidic (pH<6.3). The subpopulation of acidic phagosomes was eliminated by V-ATPase inhibition, whereas NOX inhibition caused a rapid acidification, independently of Hv1 expression. Finally, V-ATPase accumulation on phagosomes was inversely correlated to intraphagosomal ROS production in neutrophils. These data indicate that Hvcn1 ablation deregulates neutrophil pH, leading to alkalinization in phagosomes with residual ROS production or to the early accumulation of V-ATPase on phagosomes that fail to mount an oxidative response. Hv1 channels therefore differentially regulate the pH in neutrophils and macrophages, sustaining rapid acidification in macrophage phagosomes and maintaining a neutral pH in neutrophil phagosomes.

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Myeloperoxidase-dependent fluorescein chlorination by stimulated neutrophils.

Cited by 94 publications

References 31 publications

Efficient Site-Specific Prokaryotic and Eukaryotic Incorporation of Halotyrosine Amino Acids into Proteins

Efficient Site-Specific Prokaryotic and Eukaryotic Incorporation of Halotyrosine Amino Acids into Proteins

CFTR targeting during activation of human neutrophils

Hv1 proton channels differentially regulate the pH of neutrophil and macrophage phagosomes by sustaining the production of phagosomal ROS that inhibit the delivery of vacuolar ATPases

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