Site-specific 5-hydroxytryptophan incorporation into apolipoprotein A-I impairs cholesterol efflux activity and high-density lipoprotein biogenesis
| Autor: | Anthony J. DiDonato, Stanley L. Hazen, Randall A. Hughes, Jennifer A. Buffa, Ibrahim Choucair, Bruce S. Levison, Valentin Gogonea, Maryam Zamanian-Daryoush, Ying Huang, Xinmin S. Li, Joseph A. DiDonato, Andrew D. Ellington |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2020 |
| Předmět: |
0301 basic medicine
Apolipoprotein B Oxidative phosphorylation Biochemistry 5-Hydroxytryptophan 03 medical and health sciences chemistry.chemical_compound Mice High-density lipoprotein polycyclic compounds Animals Humans Molecular Biology Mice Knockout 030102 biochemistry & molecular biology biology Apolipoprotein A-I Chemistry Cholesterol Aryldialkylphosphatase Paraoxonase nutritional and metabolic diseases Molecular Bases of Disease Biological Transport Cell Biology PON1 Molecular biology 030104 developmental biology ABCA1 biology.protein Tyrosine lipids (amino acids peptides and proteins) Lipoproteins HDL Oxidation-Reduction Lipoprotein ATP Binding Cassette Transporter 1 Protein Binding |
| Zdroj: | J Biol Chem |
| Popis: | Apolipoprotein A-I (apoA-I) is the major protein constituent of high-density lipoprotein (HDL) and a target of myeloperoxidase-dependent oxidation in the artery wall. In atherosclerotic lesions, apoA-I exhibits marked oxidative modifications at multiple sites, including Trp(72). Site-specific mutagenesis studies have suggested, but have not conclusively shown, that oxidative modification of Trp(72) of apoA-I impairs many atheroprotective properties of this lipoprotein. Herein, we used genetic code expansion technology with an engineered Saccharomyces cerevisiae tryptophanyl tRNA-synthetase (Trp-RS):suppressor tRNA pair to insert the noncanonical amino acid 5-hydroxytryptophan (5-OHTrp) at position 72 in recombinant human apoA-I and confirmed site-specific incorporation utilizing MS. In functional characterization studies, 5-OHTrp(72) apoA-I (compared with WT apoA-I) exhibited reduced ABC subfamily A member 1 (ABCA1)-dependent cholesterol acceptor activity in vitro (41.73 ± 6.57% inhibition; p < 0.01). Additionally, 5-OHTrp(72) apoA-I displayed increased activation and stabilization of paraoxonase 1 (PON1) activity (μmol/min/mg) when compared with WT apoA-I and comparable PON1 activation/stabilization compared with reconstituted HDL (WT apoA-I, 1.92 ± 0.04; 5-OHTrp(72) apoA-I, 2.35 ± 0.0; and HDL, 2.33 ± 0.1; p < 0.001, p < 0.001, and p < 0.001, respectively). Following injection into apoA-I–deficient mice, 5-OHTrp(72) apoA-I reached plasma levels comparable with those of native apoA-I yet exhibited significantly reduced (48%; p < 0.01) lipidation and evidence of HDL biogenesis. Collectively, these findings unequivocally reveal that site-specific oxidative modification of apoA-I via 5-OHTrp at Trp(72) impairs cholesterol efflux and the rate-limiting step of HDL biogenesis both in vitro and in vivo. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|