scholarly journals Site-specific 5-hydroxytryptophan incorporation into apolipoprotein A-I impairs cholesterol efflux activity and high-density lipoprotein biogenesis

2020 ◽  
Vol 295 (15) ◽  
pp. 4836-4848 ◽  
Author(s):  
Maryam Zamanian-Daryoush ◽  
Valentin Gogonea ◽  
Anthony J. DiDonato ◽  
Jennifer A. Buffa ◽  
Ibrahim Choucair ◽  
...  
Keyword(s):  
High Density Lipoprotein ◽  
Cholesterol Efflux ◽  
Density Lipoprotein ◽  
Trna Synthetase ◽  
High Density ◽  
Oxidative Modification ◽  
Paraoxonase 1 ◽  
Site Specific ◽  
Apolipoprotein A

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 Trp72. Site-specific mutagenesis studies have suggested, but have not conclusively shown, that oxidative modification of Trp72 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-OHTrp72 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-OHTrp72 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-OHTrp72 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-OHTrp72 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 Trp72 impairs cholesterol efflux and the rate-limiting step of HDL biogenesis both in vitro and in vivo.

Synthesis of recombinant high density lipoprotein with apolipoprotein A-I and apolipoprotein A-V

2011 ◽  
Vol 392 (5) ◽  
Author(s):  
Xinbo Zhang ◽  
Baosheng Chen
Keyword(s):  
High Density Lipoprotein ◽  
Low Density Lipoprotein ◽  
Atherosclerotic Lesion ◽  
Density Lipoprotein ◽  
High Density ◽  
Over Expression ◽  
Apolipoprotein A ◽  
Atherosclerotic Lesion Development

Abstract It has been shown that apolipoprotein A-V (apoA-V) over-expression significantly lowers plasma triglyceride levels and decreases atherosclerotic lesion development. To assess the feasibility of recombinant high density lipoprotein (rHDL) reconstituted with apoA-V and apolipoprotein A-I (apoA-I) as a therapeutic agent for hyperlipidemic disorder and atherosclerosis, a series of rHDL were synthesized in vitro with various mass ratios of recombinant apoA-I and apoA-V. It is interesting to find that apoA-V of rHDL had no effect on lipoprotein lipase (LPL) activation in vitro and very low density lipoprotein (VLDL) clearance in HepG2 cells and in vivo. By contrast, LPL activation and VLDL clearance were inhibited by the addition of apoA-V to rHDL. Furthermore, the apoA-V of rHDL could not redistribute from rHDL to VLDL after incubation at 37°C for 30 min. These findings suggest that an increase of apoA-V in rHDL could not play a role in VLDL clearance in vitro and in vivo, which could, at least in part, attribute to the lost redistribution of apoA-V from rHDL to VLDL and LPL binding ability of apoA-V in rHDL. The therapeutic application of rHDL reconstituted with apoA-V and apoA-I might need the construction of rHDL from which apoA-V could freely redistribute to VLDL.

Impaired Antiatherogenic Functions of High-density Lipoprotein in Patients with Ankylosing Spondylitis

2015 ◽  
Vol 42 (9) ◽  
pp. 1652-1660 ◽  
Author(s):  
Christina Gkolfinopoulou ◽  
Efstratios Stratikos ◽  
Dimitris Theofilatos ◽  
Dimitris Kardassis ◽  
Paraskevi V. Voulgari ◽  
...  
Keyword(s):  
Ankylosing Spondylitis ◽  
High Density Lipoprotein ◽  
Antioxidant Capacity ◽  
Cholesterol Efflux ◽  
Density Lipoprotein ◽  
High Density ◽  
Paraoxonase 1 ◽  
Pon1 Activity ◽  
Akt Kinase ◽  
Increased Risk

Objective.Ankylosing spondylitis (AS) is a chronic inflammatory disease associated with increased risk of cardiovascular disease (CVD). High-density lipoprotein (HDL) exerts a series of antiatherogenic properties and protects from CVD. We evaluated whether HDL antiatherogenic properties are impaired in patients with AS.Methods.HDL (apoB-depleted serum) was isolated from 35 patients with AS and 35 age- and sex-matched controls. We measured the antioxidant capacity of HDL, the ability of HDL to induce cholesterol efflux, the activity of HDL-associated enzymes paraoxonase-1 (PON1) and myeloperoxidase (MPO), as well as the ability of HDL to induce Akt kinase activation.Results.HDL from patients with AS had decreased antioxidant capacity and decreased ability to promote cholesterol efflux from macrophages compared to controls. HDL-associated PON1 activity was lower and HDL-associated MPO activity higher in patients with AS compared to controls. Higher MPO activity correlated positively with lower antioxidant capacity of HDL in patients with AS. In addition, HDL from patients with AS had impaired endothelial Akt kinase activating properties that were inversely correlated with the MPO/PON1 ratio and positively correlated with the cholesterol efflux capacity of HDL.Conclusion.HDL from patients with AS displays impaired antiatherogenic properties. Attenuation of HDL properties may constitute a link between AS and CVD.

Abstract 389: Covalent Modifications of Apolipoproteins in High Density Lipoprotein by Oxidized Phospholipids Impair the Cholesterol Efflux Function of High Density Lipoprotein and are Detected in Hyperlipidemic Plasma

2016 ◽  
Vol 36 (suppl_1) ◽  
Author(s):  
Detao Gao ◽  
Lifang Zhang ◽  
Eugene Podrez
Keyword(s):  
High Density Lipoprotein ◽  
Human Plasma ◽  
Protein Function ◽  
Cholesterol Efflux ◽  
Density Lipoprotein ◽  
High Density ◽  
Oxidized Phospholipids ◽  
Protein Adduction ◽  
Covalent Modifications

Oxidized phospholipids (oxPLs) accumulate at sites of oxidative stress and contribute significantly to atherosclerosis and thrombosis. Most oxPLs have electrophilic substituents and are highly likely to form covalent adducts with proteins, thus compromising protein function. Detection of covalent interactions between oxPLs and proteins could provide important information regarding the type of proteins preferentially modified by oxPLs. However, to date, such studies are extremely limited due to significant technical challenges. We now carry out systematic studies on the protein adduction by oxPLs formed in murine and human plasma. Plasma samples were exposed to a physiologically relevant myeloperoxidase/H 2 O 2 /NO 2 – oxidizing system of phagocytes. Protein adduction by the oxPLs generated in plasma was assessed using LC-MS/MS after tryptic digestion and peptide enrichment using a novel method that we developed. We found that HDL apolipoproteins are the major targets of modification by oxPLs in both murine and human plasma. For apoA-I, the most abundant apolipoprotein in HDL, the major modification sites of oxPLs were located in the region (AA144-186), which is critical for the ABCA1 mediated cholesterol efflux to HDL. We further demonstrated that human apoA-I was also heavily crosslinked by specific oxPLs via histidine and lysine residues located in the region (AA144-186) with apoA-I, or other apolipoproteins, including apoA-II, apoA-IV and apoC-I. In vitro experiments demonstrated that oxPLs modification on lipid free apoA-I or nascent HDL (HDL3) dramatically impairs their function as cholesterol efflux mediators. Using hyperlipidemic LDLr-/- mice, we detected a crosslink adduct of apoA-II with apoE by oxPL in murine plasma. To the best of our knowledge, this is the first report for the detection of endogenous protein adducts with oxPLs.

Abstract 190: The Effect of Particle Size on Apolipoprotein A-I Exchange in Reconstituted and Human-derived High Density Lipoprotein Particles

2016 ◽  
Vol 36 (suppl_1) ◽  
Author(s):  
Mark S Borja ◽  
Lorena G Ortega-Guerrero ◽  
Michael N Oda
Keyword(s):  
Particle Size ◽  
High Density Lipoprotein ◽  
Cholesterol Efflux ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Density Lipoprotein ◽  
High Density ◽  
Size Exclusion ◽  
Paramagnetic Resonance ◽  
Size Dependent ◽  
Apolipoprotein A

Objective: The ability of high density lipoprotein (HDL) to promote cholesterol efflux from ATP binding cassette transporter A1 (ABCA1) is inversely associated with HDL particle size. The exchangeability of HDL’s primary protein constituent, apolipoprotein A-I (apoA-I), is positively correlated with cholesterol efflux capacity. However, it is unknown whether HDL particle size similarly affects the exchangeability of apoA-I. In this study, we investigated the effect of HDL particle size on the exchangeability of apoA-I on reconstituted (rHDL) and human-derived plasma HDL (pHDL) particles. Method and Results: Discoidal rHDL particles of well-defined size (17.0, 12.2, 9.6, 8.4 and 7.8 nm) were prepared from POPC and apoA-I by cholate dialysis and purified by size exclusion chromatography. Human HDL was isolated by subclass (HDL2b, 2a, 3a, 3b, 3c) from the plasma of healthy volunteers by sequential density gradient ultracentrifugation. HDL-apoA-I exchange (HAE) was measured by electron paramagnetic resonance (EPR) and non-denaturing gradient gel electrophoresis (NDGGE) following incubation with either spin-labeled or fluorescently-labeled, lipid-free apoA-I. Size-dependent effects were observed with rHDL particles, with the largest rHDL particles exhibiting 3-fold higher HAE compared to the smallest particles. In contrast, the HAE of human-derived pHDL particles was relatively constant across all subclasses at constant apoA-I concentration. Conclusions: HDL-apoA-I exchange is size-dependent in rHDL, with HAE positively associated with rHDL particle size, but it is size-independent in human pHDL. Our findings suggest that additional factors such as remodeling enzymes may facilitate apoA-I exchange from human pHDL.

Alteration of high-density lipoprotein functionality in Alzheimer’s disease patients

2017 ◽  
Vol 95 (8) ◽  
pp. 894-903 ◽  
Author(s):  
Paméla Camponova ◽  
Aurélie Le Page ◽  
Hicham Berrougui ◽  
Julie Lamoureux ◽  
Graham Pawelec ◽  
...  
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
High Density Lipoprotein ◽  
Healthy Subjects ◽  
Cholesterol Efflux ◽  
Density Lipoprotein ◽  
High Density ◽  
Elderly Subjects ◽  
The Difference

The aims of the present study were to determine whether high-density lipoprotein (HDL) functionality-mediated cholesterol efflux is altered in Alzheimer’s disease and to investigate the role and effect of amyloid-beta (Aβ) in the regulation of the anti-atherogenic activity of HDL. Eighty-seven elderly subjects were recruited, of whom 27 were healthy, 27 had mild cognitive impairment (MCI), and 33 had mild Alzheimer’s disease (mAD). Our results showed that total cholesterol levels are negatively correlated with the Mini-Mental State Examination (MMSE) score (r = –0.2602, p = 0.0182). HDL from the mAD patients was less efficient at mediating cholesterol efflux from J774 macrophages (p < 0.05) than HDL from the healthy subjects and MCI patients. While HDL from the MCI patients was also less efficient at mediating cholesterol efflux than HDL from the healthy subjects, the difference was not significant. Interestingly, the difference between the healthy subjects and the MCI and mAD patients with respect to the capacity of HDL to mediate cholesterol efflux disappeared when ATP-binding cassette transporter A1 (ABCA1)-enriched J774 macrophages were used. HDL fluidity was significantly inversely correlated with the MMSE scores (r = –0.4137, p < 0.009). In vitro measurements of cholesterol efflux using J774 macrophages showed that neither Aβ1-40nor Aβ1-42stimulate cholesterol efflux from unenriched J774 macrophages in basal or ABCA1-enriched J774 macrophages.

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