Details

Autor(en) / Beteiligte

• DiDonato, Joseph A.
• Aulak, Kulwant
• Huang, Ying
• Wagner, Matthew
• Gerstenecker, Gary
• Topbas, Celalettin
• Gogonea, Valentin
• DiDonato, Anthony J.
• Tang, W.H.Wilson
• Mehl, Ryan A.
• Fox, Paul L.
• Plow, Edward F.
• Smith, Jonathan D.
• Fisher, Edward A.
• Hazen, Stanley L.

Titel

Site-specific Nitration of Apolipoprotein A-I at Tyrosine 166 Is Both Abundant within Human Atherosclerotic Plaque and Dysfunctional

Ist Teil von

• The Journal of biological chemistry, 2014-04, Vol.289 (15), p.10276-10292

Ort / Verlag

United States: Elsevier Inc

Erscheinungsjahr

2014

Quelle

MEDLINE

Beschreibungen/Notizen

• We reported previously that apolipoprotein A-I (apoA-I) is oxidatively modified in the artery wall at tyrosine 166 (Tyr166), serving as a preferred site for post-translational modification through nitration. Recent studies, however, question the extent and functional importance of apoA-I Tyr166 nitration based upon studies of HDL-like particles recovered from atherosclerotic lesions. We developed a monoclonal antibody (mAb 4G11.2) that recognizes, in both free and HDL-bound forms, apoA-I harboring a 3-nitrotyrosine at position 166 apoA-I (NO2-Tyr166-apoA-I) to investigate the presence, distribution, and function of this modified apoA-I form in atherosclerotic and normal artery wall. We also developed recombinant apoA-I with site-specific 3-nitrotyrosine incorporation only at position 166 using an evolved orthogonal nitro-Tyr-aminoacyl-tRNA synthetase/tRNACUA pair for functional studies. Studies with mAb 4G11.2 showed that NO2-Tyr166-apoA-I was easily detected in atherosclerotic human coronary arteries and accounted for ∼8% of total apoA-I within the artery wall but was nearly undetectable (>100-fold less) in normal coronary arteries. Buoyant density ultracentrifugation analyses showed that NO2-Tyr166-apoA-I existed as a lipid-poor lipoprotein with <3% recovered within the HDL-like fraction (d = 1.063–1.21). NO2-Tyr166-apoA-I in plasma showed a similar distribution. Recovery of NO2-Tyr166-apoA-I using immobilized mAb 4G11.2 showed an apoA-I form with 88.1 ± 8.5% reduction in lecithin-cholesterol acyltransferase activity, a finding corroborated using a recombinant apoA-I specifically designed to include the unnatural amino acid exclusively at position 166. Thus, site-specific nitration of apoA-I at Tyr166 is an abundant modification within the artery wall that results in selective functional impairments. Plasma levels of this modified apoA-I form may provide insights into a pathophysiological process within the diseased artery wall. Background: The functional importance of apolipoprotein A-I (apoA-I) nitration at tyrosine 166 (Tyr166) in vivo is controversial. Results: Nitrotyrosine 166-apoA-I accounts for 8% of apoA-I within human atheroma, is not HDL-associated, and is functionally impaired. Conclusion: Buoyant density ultracentrifugation of HDL can lead to erroneous results, particularly with modified apoA-I forms. Significance: Detection and quantification of nitrotyrosine 166-apoA-I may provide insights into a pathophysiological process within the artery wall.