scholarly journals Formation of apolipoprotein-specific high-density lipoprotein particles from lipid-free apolipoproteins A-I and A-II

1999 ◽  
Vol 337 (3) ◽  
pp. 445-451 ◽  
Author(s):  
Moira A. CLAY ◽  
Daniel A. CEHIC ◽  
Diana H. PYLE ◽  
Kerry-Anne RYE ◽  
Philip J. BARTER
Keyword(s):  
High Density Lipoprotein ◽  
Sodium Oleate ◽  
Low Density Lipoprotein ◽  
Significant Proportion ◽  
Density Lipoprotein ◽  
High Density ◽  
Unesterified Cholesterol ◽  
Apolipoprotein A ◽  
Two Populations

We have shown previously that apolipoprotein A (apoA)-I-containing high-density lipoprotein (HDL) particles are formed by the conjugation of lipid-free apoA-I with lipids derived from other lipoprotein fractions in a process dependent on non-esterified fatty acids, generated by the lipolysis of very-low-density lipoprotein (VLDL) or provided exogenously. In the present study, we show that this process is also able to generate HDL particles containing apoA-II (A-II HDL) and both apoA-I and apoA-II (A-I/A-II HDL). When lipid-free apoA-II was incubated with either VLDLs and lipoprotein lipase or LDLs and sodium oleate, a significant proportion of the apoA-II was recovered in the HDL density fraction. This was associated with the formation of several populations of HDL-sized particles with pre-β2 electrophoretic mobility, which contained phospholipids and unesterified cholesterol as their main lipid constituents. When both lipid-free apoA-I and lipid-free apoA-II were incubated with LDL and sodium oleate, both apolipoproteins were recovered in HDLs that contained phospholipids and unesterified cholesterol as their main lipids. Two populations of particles had diameters of 7.4 and 10.8 nm and pre-β2-migration; there was also a population of pre-β1-migrating particles of diameter 4.7 nm. ApoA-I and apoA-II were both present in the larger HDLs, whereas only apoA-I was present in the smaller particles. Immunoaffinity chromatography on an anti-(apoA-I)–Sepharose column revealed that 10–20% of the apoA-II resided in particles that also contained apoA-I. The majority of the A-I/A-II HDL were present in a population of pre-β2 particles of 10.8 nm diameter. These results in vitro illustrate a potential mechanism for the formation of HDLs containing both apoA-I and apoA-II.

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.

scholarly journals Kinetics and mechanism of phosphatidylcholine and cholesterol exchange between chylomicrons and high-density lipoproteins

1983 ◽  
Vol 215 (2) ◽  
pp. 279-286 ◽  
Author(s):  
P M Lippiello ◽  
M Waite
Keyword(s):  
High Density Lipoprotein ◽  
Exchange Process ◽  
Significant Proportion ◽  
Density Lipoprotein ◽  
High Density ◽  
High Density Lipoproteins ◽  
Unesterified Cholesterol ◽  
Experimental Conditions ◽  
Kinetics Of

The exchange of phosphatidylcholine and unesterified cholesterol between rat mesenteric lymph chylomicrons and human high-density lipoproteins was studied in vitro by incubation of radiolabelled chylomicrons (with [N-methyl-14C]phosphatidylcholine and [7(n)-3H]cholesterol) with unlabelled high-density lipoproteins. The kinetic analysis was based on the extent of radioisotope exchange, which was determined by the proportion of label appearing in the high-density lipoprotein elution peak after rapid fractionation on analytical agarose columns. Under our experimental conditions, no net transfer of either phosphatidylcholine or cholesterol is observed. The kinetics of exchange of both phosphatidylcholine and cholesterol are biphasic. Over the first 30 min a maximum of 25% of the phosphatidylcholine and 33% of the cholesterol in chylomicrons exchanges rapidly into the high-density-lipoprotein fraction. Thereafter both lipids continue to exchange for up to 3 h at a much lower rate. For the rapid exchange process the calculated exchange rates for phosphatidylcholine and cholesterol are proportional to the concentrations of both chylomicrons and high-density lipoproteins. The second-order rate constants are (10.5 +/- 0.5) X 10(-5) microM-1 X min-1 for phosphatidylcholine and (32.1 +/- 4.5) X 10(-5) microM-1 X min-1 for cholesterol. The kinetics of the exchange process thus suggest that a significant proportion of both phosphatidylcholine and unesterified cholesterol is rapidly exchangeable between these lipoproteins, and that this exchange is mediated by a ‘bimolecular’, or collisional, mechanism.

scholarly journals High-density lipoprotein subpopulations as substrates for the transfer of cholesteryl esters to very-low-density lipoproteins

1990 ◽  
Vol 270 (2) ◽  
pp. 441-449 ◽  
Author(s):  
M A Lasunción ◽  
A Iglesias ◽  
N Skottová ◽  
E Orozco ◽  
E Herrera
Keyword(s):  
High Density Lipoprotein ◽  
Low Density Lipoprotein ◽  
Specific Radioactivity ◽  
Density Lipoprotein ◽  
Cholesteryl Oleate ◽  
High Density ◽  
Low Density ◽  
Very Low Density Lipoproteins ◽  
Apoprotein Composition

1. Human total HDL (high-density lipoprotein), HDL2 and HDL3 were labelled in vitro by incubation with lipoprotein-deficient serum (LPDS) which contained either [3H]cholesteryl oleate or [14C]cholesterol under different conditions. The lipoproteins were then subfractionated by heparin-Sepharose column chromatography, and three subfractions (A, B and C) were successively eluted from each preparation of HDL, HDL2 and HDL3. When the labelling was done at 37 degrees C for 17 h, the subfractions were homogeneously labelled with [3H]cholesteryl oleate. However, when it was performed for only 30 min at 4 degrees C, the subfractions showed marked differences in the 3H specific radioactivity, which was much higher in the C fractions than in the others. 2. 3H-labelled HDL2 and HDL3 subfractions behaved differently under the precipitant action of heparin-Mn2+; fraction C (the richest in apolipoprotein E) produced the largest amount of radioactive and chemical precipitate. More 3H radioactivity, but not the cholesterol, was precipitated from HDL2 or HDL3 by the reagent, demonstrating that 3H-labelled HDL2 and HDL3 behave like their fraction C, which becomes labelled to the highest specific radioactivity despite having the smallest mass. 3. The incubation of 3H-labelled HDL subfractions with human LPDS and very-low-density lipoprotein (VLDL) at 37 degrees C increased the quantity of 3H radioactivity that was precipitated, in proportion to the amount of VLDL present in the media. These changes were attributable to the action of cholesterol ester transfer protein, since they did not occur at 4 degrees C or when human LPDS was replaced with rat LPDS. 4. Kinetics of the transfer of HDL [3H]cholesteryl oleate to VLDL showed a greater apparent Vmax for fractions A than for fractions B from either HDL2 or HDL3, whereas the apparent Km values were very similar, which suggest that this transfer process is influenced by the apoprotein composition of the donor lipoprotein.

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.

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