Apolipoprotein A-I is a selective target for myeloperoxidase-catalyzed oxidation and functional impairment in subjects with cardiovascular disease
Lemin Zheng, … , Michael Kinter, Stanley L. Hazen
Lemin Zheng, … , Michael Kinter, Stanley L. Hazen
Published August 16, 2004
Citation Information: J Clin Invest. 2004;114(4):529-541. https://doi.org/10.1172/JCI21109.
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Article Cardiology

Apolipoprotein A-I is a selective target for myeloperoxidase-catalyzed oxidation and functional impairment in subjects with cardiovascular disease

Abstract

In recent studies we demonstrated that systemic levels of protein-bound nitrotyrosine (NO2Tyr) and myeloperoxidase (MPO), a protein that catalyzes generation of nitrating oxidants, serve as independent predictors of atherosclerotic risk, burden, and incident cardiac events. We now show both that apolipoprotein A-I (apoA-I), the primary protein constituent of HDL, is a selective target for MPO-catalyzed nitration and chlorination in vivo and that MPO-catalyzed oxidation of HDL and apoA-I results in selective inhibition in ABCA1-dependent cholesterol efflux from macrophages. Dramatic selective enrichment in NO2Tyr and chlorotyrosine (ClTyr) content within apoA-I recovered from serum and human atherosclerotic lesions is noted, and analysis of serum from sequential subjects demonstrates that the NO2Tyr and ClTyr contents of apoA-I are markedly higher in individuals with cardiovascular disease (CVD). Analysis of circulating HDL further reveals that higher NO2Tyr and ClTyr contents of the lipoprotein are each significantly associated with diminished ABCA1-dependent cholesterol efflux capacity of the lipoprotein. MPO as a likely mechanism for oxidative modification of apoA-I in vivo is apparently facilitated by MPO binding to apoA-I, as revealed by cross-immunoprecipitation studies in plasma, recovery of MPO within HDL-like particles isolated from human atheroma, and identification of a probable contact site between the apoA-I moiety of HDL and MPO. To our knowledge, the present results provide the first direct evidence for apoA-I as a selective target for MPO-catalyzed oxidative modification in human atheroma. They also suggest a potential mechanism for MPO-dependent generation of a proatherogenic dysfunctional form of HDL in vivo.

Authors

Lemin Zheng, Benedicta Nukuna, Marie-Luise Brennan, Mingjiang Sun, Marlene Goormastic, Megan Settle, Dave Schmitt, Xiaoming Fu, Leonor Thomson, Paul L. Fox, Harry Ischiropoulos, Jonathan D. Smith, Michael Kinter, Stanley L. Hazen

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Figure 5

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Demonstration of an interaction between MPO and apoA-I using hydrogen/de...
Demonstration of an interaction between MPO and apoA-I using hydrogen/deuterium exchange tandem MS. Exchangeable protons on HDL and MPO were each deuterium-labeled by mixing in D2O containing ND4OAc, pD 7.0, at room temperature for 1 hour. The deuterated HDL was combined with either deuterated MPO or additional deuterium buffer and incubated for 1 hour at room temperature to allow binding. Samples were diluted 25-fold into NH4OAc, pH 7.0, for 10 minutes for back (off) exchange before quenching by rapid cooling and addition of TFA to pH 2. Proteins were digested with immobilized pepsin, and then samples were immediately injected for analysis as described in Methods. The different spectra shown correspond to various stages in the hydrogen/deuterium exchange experiment. (A) ApoA-I isotopic clusters shown are for either the MPO-binding peptide A190–L203 (left) or a negative control peptide, L159–L170 (right). For each, spectra a and b contain peptide isotopic clusters before and after deuterium labeling, respectively. Spectra c and d contain deuterium-labeled peptide cluster after back (off) exchange with hydrogen in the absence and presence of MPO binding, respectively. The peptide isotopic cluster indicated by the asterisk represents a non–back-exchanged component of the A190–L203 isotope cluster due to inaccessibility of this region of apoA-I to solvent in the presence of MPO. Results shown are representative of 4 independent experiments. (B) Sequence confirmation of the identified peptic peptides was achieved by tandem MS. The collision-induced dissociation spectra and fragmentation analysis of the unlabeled and deuterium-labeled peptic peptide A190–L203 are illustrated.