Proteomic analysis of electronegative low-density lipoprotein

Abstract

This article is available online at http://www.jlr.org LDL, the main cholesterol carrier in plasma, is composed of approximately 75% lipid (mainly cholesterol) and 25% protein. The major protein in LDL is apolipoprotein B-100 (apoB-100), a protein of 550 kDa that accounts for more than 95% of the total protein mass in LDL ( 1 ). However, minor amounts of other apolipoproteins associated to LDL have been reported. Several of these proteins, such as apoE, apoC-III, and platelet-activating factor acetylhydrolase (PAF-AH), have important roles in LDL metabolism and modulate its atherogenicity despite their low concentration ( 2-4 ). Several studies have made a proteomic approach using 2D-electrophoresis or SELDI-based analysis to detect minor proteins in LDL ( 5-9 ), including apoE, apoC-III, apoC-II, apoA-I, apoA-IV, apoM, apoJ, serum amyloid A4 (SAA4), calgranulin A, lysozyme C, apoD, apoH, ␣ 1-antitrypsin, orosomucoid-1, paraoxonase-1, retinol binding protein, and prenylcysteine lyase-1.LDL is not a homogeneous entity but a group of particles that differs in density, size, electric charge, and composition. It is well established that small, dense LDL particles are more atherogenic than large, buoyant particles ( 10 ), and both of these subfractions differ in their protein content ( 6 ). Another property of LDL that confers enhanced atherogenicity is increased electronegative charge. Electronegative LDL [LDL( Ϫ )] is a minor subfraction of plasma LDL that is pro-infl ammatory and induces apoptosis in endothelial cells and leukocytes ( 11-13 ). Its relative proportion is increased in subjects Abstract Low density lipoprotein is a heterogeneous group of lipoproteins that differs in lipid and protein composition. One copy of apolipoprotein (apo)B accounts for over 95% of the LDL protein, but the presence of minor proteins could disturb its biological behavior. Our aim was to study the content of minor proteins in LDL subfractions separated by anion exchange chromatography. 29 July 2010. Published, JLR Papers in Press, August 10, 2010 DOI 10.1194 Abbreviations: apo, apolipoprotein; BHT, butylated hydroxytoluene; CE, cholesteryl ester; CETP, cholesteryl ester transfer protein; GGE, non-denaturing gradient gel electrophoresis; LDL(+), electropositive low-density lipoprotein; LDL( Ϫ ), electronegative low-density lipoprotein; Lp(a), lipoprotein (a); LRP2, low-density lipoprotein receptor-related protein 2; PAF-AH, platelet-activating factor-acetylhydrolase; SAA4, serum amyloid A4; TG, triglyceride; TTBS, Tweencontaining tris buffer saline. Supplementary key words apolipoproteins • atherosclerosis • modifi ed LDL This work was supported by Ministerio de Sanidad/Instituto de Salud Carlos III/FIS (ISCIII/FIS) Grants PI060500, CP040110 (S.B.), and CP060220 (JLS-Q.); and Ministerio de Educación y Ciencia Grant AP2004-1468 (C.B). The Proteomics Laboratory at Vall d'Hebron Research Institute is a member of the Instituto Nacional de Proteómica (ProteoRed) funded by Fundación Genoma España. CIBER de Diabetes y Enfermedades M...

ApoD mediates binding of HDL to LDL and to growing T24 carcinoma

“…We used a dualspecific ELISA in which antibodies to apoD bound intact apoB-containing lipoproteins equally efficient as antibodies specific for apoB. This property was unique for apoD and was not seen with other exchangeable apolipoproteins such apoE, apoH and apoJ, all of which are present in HDL and LDL/VLDL lipoprotein particles [32,33,34]. Adding detergent inhibited the apoD-mediated binding of apoB, demonstrating that changes in the LDL structure affect the formation of the complex.…”

Modified Forms of LDL in Plasma

“…The most abundant proteins in LDL(-) are apoA-I (0.15 molecules/particle of LDL(-)), apoE (0.22), apoC-III (0.37) and apoA-II (0.14), with a content 3-5 fold higher in LDL(-) than in native LDL (Bancells, 2010a). The role of these proteins is unclear but their relevance is probably low.…”

“…The role of these proteins is unclear but their relevance is probably low. However, other proteins whose absolute content in LDL(-) is lower, such as apoF (0.06 molecules/particle), apoJ (0.01) or PAF-AH (0.004) (Bancells, 2010a;, but their relative content compared with native LDL is 10, 20 and 100-fold higher, respectively, could have a much more relevant role. ApoF is the physiological inhibitor of cholesteryl ester transfer protein (Morton, 2008), a protein that regulates the catabolism of the VLDL-IDL-LDL cascade.…”

“…In native LDL, the content of non-apoB proteins is less than 1%. In contrast, LDL(-) contains up to 5% of non-apoB proteins (Bancells, 2010a;.…”

Overview of Current Proteomic Approaches for Discovery of Vascular Biomarkers of Atherosclerosis

“…Proteomic analysis, performed by LC MS/MS method, revealed the presence of 28 proteins most of which were involved in inflammation, coagulation and innate immunity, besides apolipoproteins involved in lipid metabolism. They observed that electronegative LDL, a minor subfraction of LDL fraction, has a higher content of minor proteins, especially apo F and apo J, compared to electropositive LDL (Bancells et al, 2010). Collins et al performed a proteomic analysis, applying LC MS/MS, to compare the lipoprotein-associated proteins derived from plasma and serum samples.…”