| 1. |
- Gangabadage, Chinthaka Saneth, et al.
(författare)
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Structure and dynamics of human apolipoprotein CIII
- 2008
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Ingår i: Journal of Biological Chemistry. - 0021-9258 .- 1083-351X. ; 283:25, s. 17416-17427
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Tidskriftsartikel (refereegranskat)abstract
- Human apolipoprotein CIII (apoCIII) is a surface component of chylomicrons, very low density lipoproteins, and high density lipoproteins. ApoCIII inhibits lipoprotein lipase as well as binding of lipoproteins to cell surface heparan sulfate proteoglycans and receptors. High levels of apoCIII are often correlated with elevated levels of blood lipids (hypertriglyceridemia). Here, we report the three-dimensional NMR structure and dynamics of human apo-CIII in complex with SDS micelles, mimicking its natural lipid-bound state. Thanks to residual dipolar coupling data, the first detailed view is obtained of the structure and dynamics of an intact apolipoprotein in its lipid-bound state. ApoCIII wraps around the micelle surface as a necklace of six approximately 10-residue amphipathic helices, which are curved and connected via semiflexible hinges. Three positively charged (Lys) residues line the polar faces of helices 1 and 2. Interestingly, their three-dimensional conformation is similar to that of the low density lipoprotein receptor binding motifs of apoE/B and the receptor-associated protein. At the C-terminal side of apoCIII, an array of negatively charged residues lines the polar faces of helices 4 and 5 and the adjacent flexible loop. Sequence comparison shows that this asymmetric charge distribution along the solvent-exposed face of apoCIII as well as other structural features are conserved among mammals. This structure provides a template for exploration of molecular mechanisms by which human apoCIII inhibits lipoprotein lipase and receptor binding.
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| 2. |
- Larsson, Mikael, et al.
(författare)
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Apolipoproteins C-I and C-III Inhibit Lipoprotein Lipase Activity by Displacement of the Enzyme from Lipid Droplets
- 2013
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Ingår i: Journal of Biological Chemistry. - 0021-9258 .- 1083-351X. ; 288:47, s. 33997-34008
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Tidskriftsartikel (refereegranskat)abstract
- Apolipoproteins (apo) C-I and C-III are known to inhibit lipoprotein lipase (LPL) activity, but the molecular mechanisms for this remain obscure. We present evidence that either apoC-I or apoC-III, when bound to triglyceride-rich lipoproteins, prevent binding of LPL to the lipid/water interface. This results in decreased lipolytic activity of the enzyme. Site-directed mutagenesis revealed that hydrophobic amino acid residues centrally located in the apoC-III molecule are critical for attachment to lipid emulsion particles and consequently inhibition of LPL activity. Triglyceride-rich lipoproteins stabilize LPL and protect the enzyme from inactivating factors such as angiopoietin-like protein 4 (angptl4). The addition of either apoC-I or apoC-III to triglyceride-rich particles severely diminished their protective effect on LPL and rendered the enzyme more susceptible to inactivation by angptl4. These observations were seen using chylomicrons as well as the synthetic lipid emulsion Intralipid. In the presence of the LPL activator protein apoC-II, more of apoC-I or apoC-III was needed for displacement of LPL from the lipid/water interface. In conclusion, we show that apoC-I and apoC-III inhibit lipolysis by displacing LPL from lipid emulsion particles. We also propose a role for these apolipoproteins in the irreversible inactivation of LPL by factors such as angptl4.
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| 3. |
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| 4. |
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| 5. |
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| 6. |
- Meyers, Nathan L, et al.
(författare)
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A Pressure-dependent Model for the Regulation of Lipoprotein Lipase by Apolipoprotein C-II
- 2015
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Ingår i: Journal of Biological Chemistry. - Bethesda : American Society for Biochemistry and Molecular Biology. - 0021-9258 .- 1083-351X. ; 290:29, s. 18029-18044
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Tidskriftsartikel (refereegranskat)abstract
- Apolipoprotein C-II (apoC-II) is the co-factor for lipoprotein lipase (LPL) at the surface of triacylglycerol-rich lipoproteins. LPL hydrolyzes triacylglycerol, which increases local surface pressure as surface area decreases and amphipathic products transiently accumulate at the lipoprotein surface. To understand how apoC-II adapts to these pressure changes, we characterized the behavior of apoC-II at multiple lipid/water interfaces. ApoC-II adsorption to a triacylglycerol/water interface resulted in large increases in surface pressure. ApoC-II was exchangeable at this interface and desorbed on interfacial compressions. These compressions increase surface pressure and mimic the action of LPL. Analysis of gradual compressions showed that apoC-II undergoes a two-step desorption, which indicates that lipid-bound apoC-II can exhibit at least two conformations. We characterized apoC-II at phospholipid/triacylglycerol/water interfaces, which more closely mimic lipoprotein surfaces. ApoC-II had a large exclusion pressure, similar to that of apoC-I and apoC-III. However, apoC-II desorbed at retention pressures higher than those seen with the other apoCs. This suggests that it is unlikely that apoC-I and apoC-III inhibit LPL via displacement of apoC-II from the lipoprotein surface. Upon rapid compressions and re-expansions, re-adsorption of apoC-II increased pressure by lower amounts than its initial adsorption. This indicates that apoC-II removed phospholipid from the interface upon desorption. These results suggest that apoC-II regulates the activity of LPL in a pressure-dependent manner. ApoC-II is provided as a component of triacylglycerol-rich lipoproteins and is the co-factor for LPL as pressure increases. Above its retention pressure, apoC-II desorbs and removes phospholipid. This triggers release of LPL from lipoproteins.
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| 7. |
- Nilsson, Stefan K, 1979-, et al.
(författare)
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Endocytosis of apolipoprotein A-V by members of the low density lipoprotein receptor and the VPS10p domain receptor families.
- 2008
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Ingår i: Journal of Biological Chemistry. - 0021-9258 .- 1083-351X. ; 283:38, s. 25920-25927
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Tidskriftsartikel (refereegranskat)abstract
- Apolipoprotein A-V (apoA-V) is present in low amounts in plasma and has been found to modulate triacylglycerol levels in humans and in animal models. ApoA-V displays affinity for members of the low density lipoprotein receptor (LDL-R) gene family, known as the classical lipoprotein receptors, including LRP1 and SorLA/LR11. In addition to LDL-A binding repeats, the mosaic receptor SorLA/LR11 also possesses a Vps10p domain. Here we show that apoA-V also binds to sortilin, a receptor from the Vsp10p domain gene family that lacks LDL-A repeats. Binding of apoA-V to sortilin was competed by neurotensin, a ligand that binds specifically to the Vps10p domain. To investigate the biological fate of receptor-bound apoA-V, binding experiments were conducted with cultured human embryonic kidney cells transfected with either SorLA/LR11 or sortilin. Compared with nontransfected cells, apoA-V binding to SorLA/LR11- and sortilin-expressing cells was markedly enhanced. Internalization experiments, live imaging studies, and fluorescence resonance energy transfer analyses demonstrated that labeled apoA-V was rapidly internalized, co-localized with receptors in early endosomes, and followed the receptors through endosomes to the trans-Golgi network. The observed decrease of fluorescence signal intensity as a function of time during live imaging experiments suggested ligand uncoupling in endosomes with subsequent delivery to lysosomes for degradation. This interpretation was supported by experiments with (125)I-labeled apoA-V, demonstrating clear differences in degradation between transfected and nontransfected cells. We conclude that apoA-V binds to receptors possessing LDL-A repeats and Vsp10p domains and that apoA-V is internalized into cells via these receptors. This could be a mechanism by which apoA-V modulates lipoprotein metabolism in vivo.
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| 8. |
- Reimund, Mart, et al.
(författare)
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Evidence for Two Distinct Binding Sites for Lipoprotein Lipase on Glycosylphosphatidylinositol-anchored High Density Lipoprotein-binding Protein 1 (GPIHBP1)
- 2015
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Ingår i: Journal of Biological Chemistry. - : American Society for Biochemistry and Molecular Biology. - 0021-9258 .- 1083-351X. ; 290:22, s. 13919-13934
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Tidskriftsartikel (refereegranskat)abstract
- GPIHBP1 is an endothelial membrane protein that transports lipoprotein lipase (LPL) from the subendothelial space to the luminal side of the capillary endothelium. Here, we provide evidence that two regions of GPIHBP1, the acidic N-terminal domain and the central Ly6 domain, interact with LPL as two distinct binding sites. This conclusion is based on comparative binding studies performed with a peptide corresponding to the N-terminal domain of GPIHBP1, the Ly6 domain of GPIHBP1, wild type GPIHBP1, and the Ly6 domain mutant GPIHBP1 Q114P. Although LPL and the N-terminal domain formed a tight but short lived complex, characterized by fast on-and off-rates, the complex between LPL and the Ly6 domain formed more slowly and persisted for a longer time. Unlike the interaction of LPL with the Ly6 domain, the interaction of LPL with the N-terminal domain was significantly weakened by salt. The Q114P mutant bound LPL similarly to the N-terminal domain of GPIHBP1. Heparin dissociated LPL from the N-terminal domain, and partially from wild type GPIHBP1, but was unable to elute the enzyme from the Ly6 domain. When LPL was in complex with the acidic peptide corresponding to the N-terminal domain of GPIHBP1, the enzyme retained its affinity for the Ly6 domain. Furthermore, LPL that was bound to the N-terminal domain interacted with lipoproteins, whereas LPL bound to the Ly6 domain did not. In summary, our data suggest that the two domains of GPIHBP1 interact independently with LPL and that the functionality of LPL depends on its localization on GPIHBP1.
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| 9. |
- Robal, Terje, et al.
(författare)
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Fatty acids bind tightly to the N-terminal domain of angiopoietin-like protein 4 and modulate its interaction with lipoprotein lipase
- 2012
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Ingår i: Journal of Biological Chemistry. - Bethesda : American Society for Biochemistry and Molecular Biology. - 0021-9258 .- 1083-351X. ; 287:35, s. 29739-29752
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Tidskriftsartikel (refereegranskat)abstract
- Angiopoietin-like protein 4 (Angptl4), a potent regulator of plasma triglyceride metabolism, binds to lipoprotein lipase (LPL) through its N-terminal coiled-coil domain (ccd-Angptl4) inducing dissociation of the dimeric enzyme to inactive monomers. In the present study we demonstrate that fatty acids reduce the inactivation of LPL by Angptl4. This was the case both with ccd-Angptl4 and full length Angptl4 and the effect was seen in human plasma or in the presence of albumin. The effect decreased in the sequence oleic acid > palmitic acid > myristic acid >linoleic acid >linolenic acid. Surface plasmon resonance, isothermal titration calorimetry, fluorescence and chromatography measurements revealed that fatty acids bind with high affinity to ccd-Angptl4. The interactions were characterized by fast association and slow dissociation rates, indicating formation of stable complexes. The highest affinity for ccd-Angptl4 was detected for oleic acid with a sub-nanomolar equilibrium dissociation constant (Kd). The Kd values for palmitic and myristic acid were in nanomolar range. Linoleic and linolenic acid bound with much lower affinity. On binding of fatty acids, ccd-Angptl4 underwent conformational changes resulting in a decreased helical content, weakened structural stability, dissociation of oligomers, and altered fluorescence properties of the Trp38 residue which is located close to the putative LPL-binding region. Based on these results, we propose that fatty acids play an important role in modulating the effects of Angptl4.
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| 10. |
- Shen, Yan, et al.
(författare)
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Site-directed mutagenesis of apolipoprotein CII to probe the role of its secondary structure for activation of lipoprotein lipase.
- 2010
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Ingår i: Journal of Biological Chemistry. - 0021-9258 .- 1083-351X. ; 285:10, s. 7484-7492
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Tidskriftsartikel (refereegranskat)abstract
- Apolipoprotein CII (apoCII) is a necessary activator for lipoprotein lipase (LPL). We had identified four residues (Tyr-63, Ile-66, Asp-69, and Gln-70), presumably contained in an alpha-helix, as a potential binding site for LPL. We have now used structure prediction, mutagenesis, and functional assays to explore the functional role of the secondary structure in this part of apoCII. First, mutants were generated by replacements with proline residues to disturb the helical structure. Activation by mutant G65P was reduced by 30%, whereas mutant S54P retained activation ability. Mutants V71P and L72P should be located outside the LPL-binding site, but V71P was totally inactive, whereas activation by L72P was reduced by 65%. Insertion of alanine after Tyr-63, changing the position of the putative LPL-binding site in relation to the hydrophobic face of the alpha-helix, also severely impeded the activation ability, and a double mutant (Y63A/I66A) was completely inactive. Next, to investigate the importance of conserved hydrophobic residues in the C-terminal end of apoCII, Phe-67, Val-71, Leu-72, and Leu-75 were exchanged for polar residues. Only F67S showed dramatic loss of function. Finally, fragment 39-62, previously claimed to activate LPL, was found to be completely inactive. Our data support the view that the helical structure close to the C-terminal end of apoCII is important for activation of LPL, probably by placing residues 63, 66, 69, and 70 in an optimal steric position. The structural requirements for the hydrophobic face on the back side of this helix and further out toward the C terminus were less stringent.
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