Packing of cholesterol molecules in human high-density lipoproteins

Lund‐Katz, Sissel; Phillips, Michael C.

doi:10.1021/bi00301a015

Cited by 52 publications

(39 citation statements)

References 36 publications

(43 reference statements)

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“…For TAG between the surface and core of VLDLy, ⌬v ‫ف‬ 59 Hz and the residence time of TAG at either the surface or the core is у7.5 ms. This is similar to values observed for the residence time of cholesterol in either the surface or core of HDL, у10 ms (21).…”

Section: Discussionsupporting

confidence: 86%

“…The identification of TAG in the surface of PL-stabilized emulsions indicates that TAG is likely present in the surface of native lipoproteins but is of insufficient 13 C signal intensity to be detected by NMR (17). Thus, although numerous 13 C NMR spectra of native lipoproteins have been published, surface-located TAG has not previously been detected (19)(20)(21)(22) by this approach.…”

mentioning

confidence: 92%

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Integral apolipoproteins increase surface-located triacylglycerol in intact native apoB-100-containing lipoproteins

Boyle-Roden

Walzem

2005

Journal of Lipid Research

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High-resolution NMR was used to measure the presence and quantity of triacylglycerol (TAG) in the surface of intact native apolipoprotein B-100-containing lipoprotein particles that are made by chickens in response to estrogen treatment and that in hens are deposited in yolk follicles (VLDLy). Integration of 13 C NMR resonances shows that intact VLDLy particles contain more surface TAG (5.1 ؎ 0.6 mol%, 6.7 ؎ 0.8 weight %) than predicted by apolipoprotein-free models using similarly acyl-heterogenous TAG. Lipoprotein particle metabolism exemplifies the complex biological chemistry that is reliant on the integrated function of apolipoproteins, enzymes, and transfer proteins and fundamentally dependent on the accessibility of lipoprotein lipids to those proteins. Mechanistic studies often focus on protein-based interactions that regulate particle metabolism and uptake. Tremendous advances in genetic manipulation and protein chemistry continue to drive rapid progress in these areas. Studies with model triacylglycerol (TAG)-rich lipoprotein particles show that metabolism also depends on surface lipid composition (1, 2). Paradoxical outcomes, such as increased apparent lipase activity with low-affinity substrates, occur when only whole particle lipid substrate composition is considered (2). This occurs presumably because most reactions involving nonpolar lipids take place at the interface between the particle and the surrounding aqueous medium; thus, both the affinity of the enzyme for the interface and the specific accessibility of the substrate molecules to the lipase drive reaction kinetics, rather than total particle lipid content per se (2). The presence of relatively small quantities of both substrate and nonsubstrate lipids can alter the affinity of an enzyme for the interface (3-5). Thus, the more metabolically relevant description of lipoprotein particle lipids is their specific composition and concentration within the surface layer.Lipoprotein particles possess an outer amphipathic layer of proteins, unesterified cholesterol (UC), and phospholipids (PLs) and an inner core of nonpolar TAG and cholesteryl ester (CE). The major surface components of apolipoprotein B-100 (apoB-100)-containing lipoprotein particles are well characterized in terms of their general mass, orientation, and surface area (6-9). Miller and Small (8) prepared emulsions from the lipids extracted from human VLDLs and then broke these same emulsions by ultracentrifugation to obtain a pelleted surface and a floating core, or oil, phase. Lipids present in each physically separated phase were quantitated and used to determine the distribution of lipid classes between surface and core components of these emulsions. TAG accounted for 3.63 Ϯ 0.6 weight percent (wt%) of the surface of those emulsions, whereas in a similarly prepared sample of mon-

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Section: Discussionsupporting

confidence: 86%

mentioning

confidence: 92%

Integral apolipoproteins increase surface-located triacylglycerol in intact native apoB-100-containing lipoproteins

Boyle-Roden

Walzem

2005

Journal of Lipid Research

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“…Human plasma apoA-I was purified as previously described (23). The ABCA1-GFP construct (a gift of Dr. Richard Lawn) in the mammalian expression vector pEGFP-N1 (Clontech; Palo Alto, CA) was stably expressed in CHO cells using the Lipofectamine Plus kit (Invitrogen) following the manufacturer's instructions.…”

Section: Methodsmentioning

confidence: 99%

An analysis of the role of a retroendocytosis pathway in ABCA1-mediated cholesterol efflux from macrophages

Faulkner

Panagotopulos

Johnson

et al. 2008

Journal of Lipid Research

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The ATP binding cassette transporter A-1 (ABCA1) is critical for apolipoprotein-mediated cholesterol efflux, an important mechanism employed by macrophages to avoid becoming lipid-laden foam cells, the hallmark of early atherosclerotic lesions. It has been proposed that lipid-free apolipoprotein A-I (apoA-I) enters the cell and is resecreted as a lipidated particle via a retroendocytosis pathway during ABCA1-mediated cholesterol efflux from macrophages. To determine the functional importance of such a pathway, confocal microscopy was used to characterize the internalization of a fully functional apoA-I cysteine mutant containing a thiol-reactive fluorescent probe in cultured macrophages. ApoA-I was also endogenously labeled with 35 S-methionine to quantify cellular uptake and to determine the metabolic fate of the internalized protein. It was found that apoA-I was specifically taken inside macrophages and that a small amount of intact apoA-I was resecreted from the cells. However, a majority of the label that reappeared in the media was degraded. We estimate that the mass of apoA-I retroendocytosed is not sufficient to account for the HDL produced by the cholesterol efflux reaction. Furthermore, we have demonstrated that lipid-free apoA-Imediated cholesterol efflux from macrophages can be pharmacologically uncoupled from apoA-I internalization into cells. On the basis these findings, we present a model in which the ABCA1-mediated lipid transfer process occurs primarily at the membrane surface in macrophages, but still accounts for the observed specific internalization of apoA-I.-

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“…Low density and high density lipoproteins (LDL and HDL) were prepared from fresh human plasma by ultracentrifugation (Hatch and Lees, 1968). Egg PC small unilamellar vesicles (PC-SUV) and reconstituted PC-apoHDL discs (PC/protein ratio ϭ 2.1, w/w) were prepared by sonication (Barenholz et al, 1977;Lund-Katz and Phillips, 1984). Acceptor preparations were dialyzed against the appropriate tissue culture medium and filter sterilized (0.45-m pore size) before use.…”

Section: Methodsmentioning

confidence: 99%

Efflux of Newly Synthesized Cholesterol and Biosynthetic Sterol Intermediates from Cells

Johnson

Fischer

Phillips

et al. 1995

Journal of Biological Chemistry

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Previous studies suggest that during sterol synthesis in cells, cholesterol and precusor sterols are transported to the plasma membrane and that this transport is stimulated by the binding of high density lipoprotein (HDL) to its putative cell surface receptor, leading to enhanced sterol efflux. Little is known about the identities of synthesized sterols subject to efflux or whether efflux of cholesterol and precursor sterols are stimulated equally by HDL. To address these issues, cells were incubated with [ 3 H]acetate or [ 3 H]mevalonate and sterol acceptors, and then the labeled sterols in cells and efflux media were analyzed by high pressure liquid chromatography methods that resolved cholesterol and precursor sterols. In non-hepatic cells (Chinese hamster ovary (CHO), fibroblasts, and smooth muscle), cholesterol and multiple precursor sterols accumulated. In CHO cells, the major products were cholesterol and desmosterol, which together constituted 50% of labeled nonsaponifiable lipids. When media contained human HDL 3 (1 mg of protein/ml), the molar efflux of synthesized desmosterol was four times that of cholesterol, and the 8-h efflux of these sterols, each normalized to its own production, averaged 48 and 16%, respectively. When media contained egg phosphatidylcholine vesicles (1 mg/ml), the efflux of these sterols averaged 18 and 2.4%, respectively. Thus, with both acceptors, desmosterol was the major synthesized sterol released from cells, and its efflux was substantially greater than that of synthesized cholesterol. High relative efflux of desmosterol (or a desmosterol-like sterol) occurred in all cell types and in both cholesterol-enriched and unenriched cells. These results demonstrated qualitatively similar efflux of synthesized sterols in the presence of HDL 3 and phospholipid vesicles, arguing against an absolute requirement for acceptors that interact with the HDL receptor. To probe for possible quantitative differences in the capabilities of these two acceptors, the ratios of (efflux to HDL 3 )/(efflux to phosphatidylcholine vesicles) were calculated for synthesized cholesterol and desmosterol, plasma membrane cholesterol, and lysosomal cholesterol. In comparison to plasma membrane cholesterol, there was little or no HDL selectivity for lysosomal cholesterol or synthesized desmosterol, whereas there was a 2-3-fold selectivity for synthesized cholesterol, suggesting that the ability of HDL to enhance the efflux of synthesized sterols is a modest quantitative effect and confined to cholesterol.

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Packing of cholesterol molecules in human high-density lipoproteins

Cited by 52 publications

References 36 publications

Integral apolipoproteins increase surface-located triacylglycerol in intact native apoB-100-containing lipoproteins

Integral apolipoproteins increase surface-located triacylglycerol in intact native apoB-100-containing lipoproteins

An analysis of the role of a retroendocytosis pathway in ABCA1-mediated cholesterol efflux from macrophages

Efflux of Newly Synthesized Cholesterol and Biosynthetic Sterol Intermediates from Cells

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