title: Tyrosine 192 in apolipoprotein A-I is the major site of nitration and chlorination by myeloperoxidase, but only chlorination markedly impairs ABCA1-dependent cholesterol transport.
abstract: High density lipoprotein (HDL) isolated from human atherosclerotic lesions and the blood of patients with established coronary artery disease contains elevated levels of 3-nitrotyrosine and 3-chlorotyrosine. Myeloperoxidase (MPO) is the only known source of 3-chlorotyrosine in humans, indicating that MPO oxidizes HDL in vivo. In the current studies, we used tandem mass spectrometry to identify the major sites of tyrosine oxidation when lipid-free apolipoprotein A-I (apoA-I), the major protein of HDL, was exposed to MPO or peroxynitrite (ONOO(-)). Tyrosine 192 was the predominant site of both nitration and chlorination by MPO and was also the major site of nitration by ONOO(-). Electron paramagnetic spin resonance studies of spin-labeled apoA-I revealed that residue 192 was located in an unusually hydrophilic environment. Moreover, the environment of residue 192 became much more hydrophobic when apoA-I was incorporated into discoidal HDL, and Tyr(192) of HDL-associated apoA-I was a poor substrate for nitration by both myeloperoxidase and ONOO(-), suggesting that solvent accessibility accounted in part for the reactivity of Tyr(192). The ability of lipid-free apoA-I to facilitate ATP-binding cassette transporter A1 cholesterol transport was greatly reduced after chlorination by MPO. Loss of activity occurred in concert with chlorination of Tyr(192). Both ONOO(-) and MPO nitrated Tyr(192) in high yield, but unlike chlorination, nitration minimally affected the ability of apoA-I to promote cholesterol efflux from cells. Our results indicate that Tyr(192) is the predominant site of nitration and chlorination when MPO or ONOO(-) oxidizes lipid-free apoA-I but that only chlorination markedly reduces the cholesterol efflux activity of apoA-I. This impaired biological activity of chlorinated apoA-I suggests that MPO-mediated oxidation of HDL might contribute to the link between inflammation and cardiovascular disease.
authors.affiliation: Department of Medicine, University of Washington, Seattle, WA 98195, USA.
authors.forename: Baohai
authors.initials: B
authors.lastname: Shao
authors.forename: Constanze
authors.initials: C
authors.lastname: Bergt
authors.forename: Xiaoyun
authors.initials: X
authors.lastname: Fu
authors.forename: Pattie
authors.initials: P
authors.lastname: Green
authors.forename: John C
authors.initials: JC
authors.lastname: Voss
authors.forename: Michael N
authors.initials: MN
authors.lastname: Oda
authors.forename: John F
authors.initials: JF
authors.lastname: Oram
authors.forename: Jay W
authors.initials: JW
authors.lastname: Heinecke
mesh_terms: D064286:ATP Binding Cassette Transporter 1; D018528:ATP-Binding Cassette Transporters / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D000975:Antioxidants / Q000378:metabolism / Q000494:pharmacology; D016632:Apolipoprotein A-I / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001692:Biological Transport / Q000187:drug effects; D002460:Cell Line; D002713:Chlorine / Q000378:metabolism*; D002784:Cholesterol / Q000378:metabolism*; D006224:Cricetinae; D004578:Electron Spin Resonance Spectroscopy; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism; D057927:Hydrophobic and Hydrophilic Interactions; D009566:Nitrates / Q000378:metabolism* / Q000494:pharmacology; D009584:Nitrogen / Q000378:metabolism; D010084:Oxidation-Reduction; D010455:Peptides / Q000737:chemistry / Q000378:metabolism; D009195:Peroxidase / Q000378:metabolism*; D030421:Peroxynitrous Acid / Q000378:metabolism / Q000494:pharmacology; D021241:Spectrometry, Mass, Electrospray Ionization; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000235:genetics / Q000378:metabolism*
chemical_list: C577836:ABCA1 protein, human; D064286:ATP Binding Cassette Transporter 1; D018528:ATP-Binding Cassette Transporters; D000975:Antioxidants; D016632:Apolipoprotein A-I; D009566:Nitrates; D010455:Peptides; D030421:Peroxynitrous Acid; C002744:3-nitrotyrosine; D014443:Tyrosine; D002713:Chlorine; D002784:Cholesterol; D006861:Hydrogen Peroxide; D009195:Peroxidase; D009584:Nitrogen