The Structure of Apolipoprotein A-II in Discoidal High Density Lipoproteins

Silva, R.A. Gangani D.; Schneeweis, Lumelle A.; Krishnan, Srinivasan; Zhang, Xiuqi; Axelsen, Paul H.; Davidson, W. Sean

doi:10.1074/jbc.m610380200

Cited by 35 publications

(52 citation statements)

References 34 publications

(53 reference statements)

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“…Monomeric apoA-IV was exhaustively digested with sequencing grade trypsin (5% (w/w)) overnight, and 75-100-g aliquots of trypsinized cross-linked monomeric apoA-IV were lyophilized and stored at Ϫ20°C until use. Digested protein samples were analyzed on an Applied Biosystems Q-Star XL mass spectrometer equipped with an online Agilent 1100 capillary high performance liquid chromatograph as described previously (31,32). The crosslinked peptide pair described in this paper (1-25 ϫ 333-343) was identified by mass and by tandem mass spectrometry.…”

Section: Methodsmentioning

confidence: 99%

Modulation of Apolipoprotein A-IV Lipid Binding by an Interaction between the N and C Termini

Tubb

Silva

Tso

et al. 2007

Journal of Biological Chemistry

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Apolipoprotein A-IV (apoA-IV) is a 376-amino acid exchangeable apolipoprotein made in the small intestine of humans. Although it has many proposed roles in vascular disease, satiety, and chylomicron metabolism, there is no known structural basis for these functions. The ability to associate with lipids may be a key step in apoA-IV functionality. We recently identified a single amino acid, Phe 334 , which seems to inhibit the lipid binding capability of apoA-IV. We also found that an intact N terminus was necessary for increased lipid binding of Phe 334 mutants. Here, we identify Trp 12 and Phe 15 as the N-terminal amino acids required for the fast lipid binding seen with the F334A mutant. Furthermore, we found that individual disruption of putative amphipathic ␣-helices 3-11 had little effect on lipid binding, suggesting that the N terminus of apoA-IV may be the operational site for initial lipid binding. We also provide three independent pieces of experimental evidence supporting a direct intramolecular interaction between sequences near amino acids 12/15 and 334. This interaction could represent a unique "switch" mechanism by which apoA-IV changes lipid avidity in vivo.

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Section: Methodsmentioning

confidence: 99%

Modulation of Apolipoprotein A-IV Lipid Binding by an Interaction between the N and C Termini

Tubb

Silva

Tso

et al. 2007

Journal of Biological Chemistry

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“…HDLs used in the study were reconstituted using either spontaneous solubilization of multilamellar vesicles or by Biobead-cholate removal method (9,20,24). See supplementary data for detailed information on different types of particle preparations.…”

Section: Preparation Of Rhdlmentioning

confidence: 99%

“…Cross-linking of the particles was carried out as essentially described before (9). See supplementary data for details on the cross-linking procedure.…”

Section: Cross-linking Of Rhdl Particlesmentioning

confidence: 99%

“…Structural studies on similar diameter reconstituted discoidal (2 apoA-I/particle) and spherical (3 apoA-I/particle) HDL particles have indicated that apoA-I adopts a common organizational motif, i.e., "double belt" model, irrespective of particle shape, size, and the number of apoA-I molecules in the particle (8). In this model, two apoA-I molecules are arranged in an antiparallel fashion forming a double belt that is stabilized by salt bridge interactions (9,10). The interaction of lipid-free apoA-I with ABCA1-expressing cells generates discoidal nascent HDL with two to four molecules of apoA-I/ particle (11)(12)(13).…”

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confidence: 99%

“…The apolipoprotein/particle stoichiometry is determined by dividing the estimated M r of the cross-linked complex by the actual M r of monomeric apoA-I. However, at low protein:cross-linker ratio, even for rHDL with two molecules of apoA-I, one sees multiple SDS-PAGGE bands due to the extent and sequence location of the intermolecular cross-links (9,19). At high protein:cross-linker ratio, band smearing of the cross-linked complex is common, and hence the migration distance of the band is difficult to measure.…”

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Mass spectrometric determination of apolipoprotein molecular stoichiometry in reconstituted high density lipoprotein particles

Massey

Pownall

Macha

et al. 2009

Journal of Lipid Research

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Plasma HDL-cholesterol and apolipoprotein A-I (apoA-I) levels are strongly inversely associated with cardiovascular disease. However, the structure and protein composition of HDL particles is complex, as native and synthetic discoidal and spherical HDL particles can have from two to five apoA-I molecules per particle. To fully understand structure-function relationships of HDL, a method is required that is capable of directly determining the number of apolipoprotein molecules in heterogeneous HDL particles. Chemical cross-linking followed by SDS polyacrylamide gradient gel electrophoresis has been previously used to determine apolipoprotein stoichiometry in HDL particles. However, this method yields ambiguous results due to effects of cross-linking on protein conformation and, subsequently, its migration pattern on the gel. Here, we describe a new method based on cross-linking chemistry followed by MALDI mass spectrometry that determines the absolute mass of the cross-linked complex, thereby correctly determining the number of apolipoprotein molecules in a given HDL particle. Using well-defined, homogeneous, reconstituted apoA-I-containing HDL, apoA-IV-containing HDL, as well as apoA-I/apoA-II-containing HDL, we have validated this method. The method has the capability to determine the molecular ratio and molecular composition of apolipoprotein molecules in complex reconstituted HDL

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Apolipoprotein Gene Structure and Function

Martín‐Campos

Chang

Blanco‐Vaca

et al. 2017

Encyclopedia of Life Sciences

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Plasma lipoproteins are lipid–protein complexes that circulate in blood. The protein components in these complexes are called apolipoproteins. The genes for the ‘soluble’ apolipoproteins, that is apoA1, A2, A4, A5, C1, C2, C3 and apoE, are structurally and evolutionarily related; they display similar structures that include four exons and three introns in specific locations (except A4, which contains only two introns, missing intron 1 in the 5′‐untranslated region). The proteins contain internal repeats of well‐defined amphipathic lipid‐binding motifs. ApoB‐100 is an ‘insoluble’ apolipoprotein in low‐density lipoprotein. ApoB mRNA editing in the small intestine generates a premature stop codon; translation of the edited mRNA produces apoB‐48, a protein ∼48% the size of apoB‐100 that is required for the biogenesis of chylomicrons. Apo(a) is a large glycoprotein which is covalently linked to apoB‐100 by a disulfide bond. Such apo B‐100‐apo(a) complex is a distinguishing feature of lipoprotein(a) (Lp(a)). In addition to the classic apolipoproteins, a host of other proteins have been found in lipoproteins. These proteins are often better known for their nonlipoprotein‐related functions, and their genes are structurally unrelated to those of the classic apolipoproteins. Many of the gene products discussed in this article play important roles in disease pathogenesis, diagnosis, prognostication and therapy. Key Concepts Apolipoproteins are the main protein components of lipoproteins. Apolipoproteins display other specialised functions in addition to their role as lipid carriers. A few apolipoproteins have been found to be predictors of neurodegenerative diseases or as potential prognostic markers of several types of cancers. Some apolipoproteins are modulators of both innate and adaptive immune responses. The ApoA1/C3/A4 gene cluster plays multiple pathophysiologic roles in lipoprotein metabolism and maintenance of plasma lipid levels and, as such, may influence the risk of cardiovascular disease development. ApoB‐100, the major apolipoprotein in LDL and VLDL, and apoB‐48, an essential apolipoprotein in chylomicron biogenesis, have structural elements that are very different from those of the soluble apolipoproteins. Studies on the biogenesis of apoB‐48 led to the discovery of eukaryotic RNA editing. Plasma levels of Lp(a) are a major independent risk indicator of premature coronary artery disease and stroke.

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The Structure of Apolipoprotein A-II in Discoidal High Density Lipoproteins

Cited by 35 publications

References 34 publications

Modulation of Apolipoprotein A-IV Lipid Binding by an Interaction between the N and C Termini

Modulation of Apolipoprotein A-IV Lipid Binding by an Interaction between the N and C Termini

Mass spectrometric determination of apolipoprotein molecular stoichiometry in reconstituted high density lipoprotein particles

Apolipoprotein Gene Structure and Function

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