Human Apolipoprotein Research Articles

Quantification by nano liquid chromatography parallel reaction monitoring mass spectrometry of human apolipoprotein A-I, apolipoprotein B, and hemoglobin A1c in dried blood spots.

Proteomic analysis of blood proteins in dried blood spots (DBS) is gaining attention as a possible replacement for measurements in plasma/serum collected by venipuncture. We aimed to develop and provisionally validate a nanoflow LC-PRM-MS method for clinical use. We used Skyline to develop a nanoflow LC-PRM-MS method to quantify glycated hemoglobin-β, apolipoprotein A-I, and apolipoprotein B in DBS. Precision, linearity, interferences, and stability were determined and the method was used to analyze samples from 36 human volunteers. The method was compared with clinically validated measurements in paired blood collected via venipuncture. The method was relatively precise for these proteins (10-11% CV) and linear across the normal concentration ranges of these proteins. Interference from high total serum protein concentration (>8 g/dL) was noted for apolipoprotein A-I and apolipoprotein B. Proteins in DBS were stable for 14 days at temperatures below 25°C and trypsinized samples were stable for 48 h at 7°C. There was moderate correlation with clinical methods (r = 0.783-0.858) and significant bias in individual samples. Although the method had adequate precision and linearity for a biomarker, the accuracy compared with clinically validated assays raises concerns regarding the use of DBS compared with venipuncture for clinical use.

Liver X receptor agonist treatment significantly affects phenotype and transcriptome of APOE3 and APOE4 Abca1 haplo-deficient mice.

ATP-binding cassette transporter A1 (ABCA1) controls cholesterol and phospholipid efflux to lipid-poor apolipoprotein E (APOE) and is transcriptionally controlled by Liver X receptors (LXRs) and Retinoic X Receptors (RXRs). In APP transgenic mice, lack of Abca1 increased Aβ deposition and cognitive deficits. Abca1 haplo-deficiency in mice expressing human APOE isoforms, increased level of Aβ oligomers and worsened memory deficits, preferentially in APOE4 mice. In contrast upregulation of Abca1 by LXR/RXR agonists significantly ameliorated pathological phenotype of those mice. The goal of this study was to examine the effect of LXR agonist T0901317 (T0) on the phenotype and brain transcriptome of APP/E3 and APP/E4 Abca1 haplo-deficient (APP/E3/Abca1+/- and APP/E4/Abca1+/-) mice. Our data demonstrate that activated LXRs/RXR ameliorated APOE4-driven pathological phenotype and significantly affected brain transcriptome. We show that in mice expressing either APOE isoform, T0 treatment increased mRNA level of genes known to affect brain APOE lipidation such as Abca1 and Abcg1. In both APP/E3/Abca1+/- and APP/E4/Abca1+/- mice, the application of LXR agonist significantly increased ABCA1 protein level accompanied by an increased APOE lipidation, and was associated with restoration of APOE4 cognitive deficits, reduced levels of Aβ oligomers, but unchanged amyloid load. Finally, using Gene set enrichment analysis we show a significant APOE isoform specific response to LXR agonist treatment: Gene Ontology categories “Microtubule Based Process” and “Synapse Organization” were differentially affected in T0-treated APP/E4/Abca1+/- mice. Altogether, the results are suggesting that treatment of APP/E4/Abca1+/- mice with LXR agonist T0 ameliorates APOE4-induced AD-like pathology and therefore targeting the LXR-ABCA1-APOE regulatory axis could be effective as a potential therapeutic approach in AD patients, carriers of APOEε4.

Oxidative stress-mediated NFκB phosphorylation upregulates p62/SQSTM1 and promotes retinal pigmented epithelial cell survival through increased autophagy.

p62 is a scaffolding adaptor implicated in the clearance of protein aggregates by autophagy. Reactive oxygen species (ROS) can either stimulate or inhibit NFκB-mediated gene expression influencing cellular fate. We studied the effect of hydrogen peroxide (H2O2)-mediated oxidative stress and NFκB signaling on p62 expression in the retinal pigment epithelium (RPE) and investigated its role in regulation of autophagy and RPE survival against oxidative damage. Cultured human RPE cell line ARPE-19 and primary human adult and fetal RPE cells were exposed to H2O2-induced oxidative stress. The human apolipoprotein E4 targeted-replacement (APOE4) mouse model of AMD was used to study expression of p62 and other autophagy proteins in the retina. p62, NFκB p65 (total, phosphorylated, nuclear and cytoplasmic) and ATG10 expression was assessed by mRNA and protein analyses. Cellular ROS and mitochondrial superoxide were measured by CM-H2DCFDA and MitoSOX staining respectively. Mitochondrial viability was determined using MTT activity. qPCR-array system was used to investigate autophagic genes affected by p62. Nuclear and cytoplasmic levels of NFκB p65 were evaluated after cellular fractionation by Western blotting. We report that p62 is up-regulated in RPE cells under H2O2-induced oxidative stress and promotes autophagic activity. Depletion of endogenous p62 reduces autophagy by downregulation of ATG10 rendering RPE more susceptible to oxidative damage. NFκB p65 phosphorylation at Ser-536 was found to be critical for p62 upregulation in response to oxidative stress. Proteasome inhibition by H2O2 causes p62-NFκB signaling as antioxidant pre-treatment reversed p62 expression and p65 phosphorylation when RPE was challenged by H2O2 but not when by Lactacystin. p62 protein but not RNA levels are elevated in APOE4-HFC AMD mouse model, suggesting reduction of autophagic flux in disease conditions. Our findings suggest that p62 is necessary for RPE cytoprotection under oxidative stress and functions, in part, by modulating ATG10 expression. NFκB p65 activity may be a critical upstream initiator of p62 expression in RPE cells under oxidative stress.

Vaccination against T-cell epitopes of native ApoB100 reduces vascular inflammation and disease in a humanized mouse model of atherosclerosis.

The T-cell response to low-density lipoprotein (LDL) in the vessel wall plays a critical role in atherosclerotic plaque formation and stability. In this study, we used a new translational approach to investigate epitopes from human apolipoprotein B100 (ApoB100), the protein component of LDL, which triggers T-cell activation. We also evaluated the potential of two selected native ApoB100 epitopes to modulate atherosclerosis in human ApoB100-transgenic Ldlr-/- (HuBL) mice. HuBL mice were immunized with human atherosclerotic plaque homogenate to boost cellular autoimmune response to tissue-derived ApoB100 epitopes. In vitro challenge of splenocytes from immunized mice with a library of overlapping native peptides covering human ApoB100 revealed several sequences eliciting T-cell proliferation. Of these sequences, peptide (P) 265 and P295 were predicted to bind several human leucocyte antigen (HLA) haplotypes and induced high levels of interferon (IFN)-γ. Vaccination of HuBL mice with these peptides mounted a strong adaptive immune response to native ApoB100, including high levels of epitope-specific plasma IgGs. Interestingly, P265 and P295 vaccines significantly decreased plaque size, reduced macrophage infiltration and increased IgG1 deposition in the plaques. Purified IgGs from vaccinated mice displayed anti-inflammatory properties against macrophages invitro, reducing their response to LPS in a dose-dependent manner. We identified two specific epitopes from human native ApoB100 that trigger T-cell activation and protect HuBL mice against atherosclerosis when used in a vaccine. Our data suggest that vaccination-induced protective mechanisms may be mediated at least in part through specific antibody responses to LDL that inhibit macrophage activation.

Gene-environment interaction between lead and Apolipoprotein E4 causes cognitive behavior deficits in mice

BackgroundAlzheimer’s disease (AD) is characterized by progressive cognitive decline and memory loss. Environmental factors and gene-environment interactions (GXE) may increase AD risk, accelerate cognitive decline, and impair learning and memory. However, there is currently little direct evidence supporting this hypothesis.MethodsIn this study, we assessed for a GXE between lead and ApoE4 on cognitive behavior using transgenic knock-in (KI) mice that express the human Apolipoprotein E4 allele (ApoE4-KI) or Apolipoprotein E3 allele (ApoE3-KI). We exposed 8-week-old male and female ApoE3-KI and ApoE4-KI mice to 0.2% lead acetate via drinking water for 12 weeks and assessed for cognitive behavior deficits during and after the lead exposure. In addition, we exposed a second (cellular) cohort of animals to lead and assessed for changes in adult hippocampal neurogenesis as a potential underlying mechanism for lead-induced learning and memory deficits.ResultsIn the behavior cohort, we found that lead reduced contextual fear memory in all animals; however, this decrease was greatest and statistically significant only in lead-treated ApoE4-KI females. Similarly, only lead-treated ApoE4-KI females exhibited a significant decrease in spontaneous alternation in the T-maze. Furthermore, all lead-treated animals developed persistent spatial working memory deficits in the novel object location test, and this deficit manifested earlier in ApoE4-KI mice, with female ApoE4-KI mice exhibiting the earliest deficit onset. In the cellular cohort, we observed that the maturation, differentiation, and dendritic development of adult-born neurons in the hippocampus was selectively impaired in lead-treated female ApoE4-KI mice.ConclusionsThese data suggest that GXE between ApoE4 and lead exposure may contribute to cognitive impairment and that impaired adult hippocampal neurogenesis may contribute to these deficits in cognitive behavior. Together, these data suggest a role for GXE and sex differences in AD risk.

Shark-Derived Single Domain Antibodies Targeting Apolipoprotein E

Human apolipoprotein E (ApoE) exists as three common isoforms that differ by single amino acid substitutions. ApoE4 (Arg112/Arg158) is the strongest genetic risk factor for late-onset Alzheimer's disease (LOAD)—a significant cause of mortality and disability worldwide. At the protein level, previous studies suggest that the conformational landscape of ApoE4 is different to that of the wild-type ApoE3 (Cys112/Arg158). To date, however, effective ligands capable of selectively targeting the “ApoE4-conformation” remain elusive.

Human apolipoprotein A-I exerts a prophylactic effect on high-fat diet-induced atherosclerosis via inflammation inhibition in a rabbit model.

Apolipoprotein A-I (apoA-I) is the major functional protein fraction of high-density lipoprotein. The prophylactic effect and mechanism of human apoA-I on atherosclerosis (AS) were investigated in a high-fat diet-induced AS rabbit model. The rabbits were injected with apoA-I once a week while fed high-fat diet for 20 weeks. Our results showed that apoA-I could raise the serum level of high-density lipoprotein-cholesterol and reduce those of lipid total cholesterol, triglyceride, and low-density lipoprotein-cholesterol in AS rabbits. Decreased aortic plaque area and aortic injury degree were also observed by Oil Red O staining and HE staining in apoA-I-treated high-fat diet-induced AS rabbits. Further study elucidated that apoA-I could down-regulate the expression of some inflammatory mediators including intercellular adhesion molecule type 1, vascular adhesion molecule-1 (VCAM-1), monocyte chemoattractant protein-1, tumor necrosis factor-α, interleukin-6 (IL-6), and C-reactive protein in serum and aorta of AS rabbits. In addition, real-time quantitative RT-PCR analyses showed that the apoA-I infusions decreased the mRNA levels of two pro-inflammatory molecules, i.e. nuclear factor kappa B (NF-κB) and cyclooxygenase-2 (COX-2), in aorta of AS rabbits, which was associated with a concomitant reduction in endothelial VCAM-1 and IL-6 mRNA transcription. Together, our results support the atheroprotective and prophylactic role of apoA-I in vivo, and this activity may be correlated with its anti-inflammatory effect.

Chimera of Apolipophorin III and C-terminal Domain of Apolipoprotein E to Study Apolipoprotein Structure Function

Human apolipoprotein E (apoE) is a two-domain anti-atherogenic lipid transport protein that mediates plasma cholesterol homeostasis by serving as a ligand for the low density lipoprotein (LDL) receptor. Of its three isoforms E3, E2, E4, the latter two have been associated with increased risk of cardiovascular and Alzheimer's disease respectively, yet all three contain an identical C-terminal (CT) domain. The CT domain (residues 201-299) is responsible for tetramerization and has been found to initiate lipid binding that influences critical functional features of apoE. The N-terminal (NT) domain of apoE has four α-helices arranged in a bundle similar to apolipophorin III (apoLp-III), a model insect apolipoprotein containing five α-helices. To better understand the role apolipoprotein domains play in structure and function, a novel chimeric apolipoprotein was designed by attaching apoE-CT to apoLp-III. Recombinant apoLp-III/apoE-CT, apoE3, apoE-CT, and apoLp-III were expressed in bacterial cells, purified by affinity chromatography, and purity verified by SDS-PAGE. Western blot analysis using monoclonal apoE-CT specific antibody confirmed the presence of apoE-CT in the chimera. Crosslinking studies using dimethylsuberimidate revealed that the apoLp-III/apoE-CT chimera formed oligomers similar to apoE, while apoLp-III was monomeric. The chimera was highly α-helical, and incubation with 1-anilinonaphthalene-8-sulfonic acid showed increased fluorescence intensity when apoE-CT was added to apoLp-III, consistent with the presence of water-shielded dye binding sites upon addition of a structured apoE-CT segment. Our results suggest that the chimeric protein has structural features similar to the parent proteins, and that self-association properties of apoE can be transferred to apoLp-III. Future studies will include measuring the ability of the chimera to solubilize phospholipid bilayers and binding to low density lipoproteins. The chimeric approach offers the potential to obtain insight into domain interactions and the structure-function relationships in apolipoproteins.

Novel human bioactive peptides identified in Apolipoprotein B: Evaluation of their therapeutic potential

Host defence peptides (HDPs) are short, cationic amphipathic peptides that play a key role in the response to infection and inflammation in all complex life forms. It is increasingly emerging that HDPs generally have a modest direct activity against a broad range of microorganisms, and that their anti-infective properties are mainly due to their ability to modulate the immune response. Here, we report the recombinant production and characterization of two novel HDPs identified in human Apolipoprotein B (residues 887–922) by using a bioinformatics method recently developed by our group. We focused our attention on two variants of the identified HDP, here named r(P)ApoBL and r(P)ApoBS, 38- and 26-residue long, respectively. Both HDPs were found to be endowed with a broad-spectrum antimicrobial activity while they show neither toxic nor haemolytic effects towards eukaryotic cells. Interestingly, both HDPs were found to display a significant anti-biofilm activity, and to act in synergy with either commonly used antibiotics or EDTA. The latter was selected for its ability to affect bacterial outer membrane permeability, and to sensitize bacteria to several antibiotics. Circular dichroism analyses showed that SDS, TFE, and LPS significantly alter r(P)ApoBL conformation, whereas slighter or no significant effects were detected in the case of r(P)ApoBS peptide. Interestingly, both ApoB derived peptides were found to elicit anti-inflammatory effects, being able to mitigate the production of pro-inflammatory interleukin-6 and nitric oxide in LPS induced murine macrophages. It should also be emphasized that r(P)ApoBL peptide was found to play a role in human keratinocytes wound closure in vitro. Altogether, these findings open interesting perspectives on the therapeutic use of the herein identified HDPs.

Helical structure, stability, and dynamics in human apolipoprotein E3 and E4 by hydrogen exchange and mass spectrometry

Apolipoprotein E (apoE) plays a critical role in cholesterol transport in both peripheral circulation and brain. Human apoE is a polymorphic 299-residue protein in which the less common E4 isoform differs from the major E3 isoform only by a C112R substitution. ApoE4 interacts with lipoprotein particles and with the amyloid-β peptide, and it is associated with increased incidence of cardiovascular and Alzheimer's disease. To understand the structural basis for the differences between apoE3 and E4 functionality, we used hydrogen-deuterium exchange coupled with a fragment separation method and mass spectrometric analysis to compare their secondary structures at near amino acid resolution. We determined the positions, dynamics, and stabilities of the helical segments in these two proteins, in their normal tetrameric state and in mutation-induced monomeric mutants. Consistent with prior X-ray crystallography and NMR results, the N-terminal domain contains four α-helices, 20 to 30 amino acids long. The C-terminal domain is relatively unstructured in the monomeric state but forms an α-helix ∼70 residues long in the self-associated tetrameric state. Helix stabilities are relatively low, 4 kcal/mol to 5 kcal/mol, consistent with flexibility and facile reversible unfolding. Secondary structure in the tetrameric apoE3 and E4 isoforms is similar except that some helical segments in apoE4 spanning residues 12 to 20 and 204 to 210 are unfolded. These conformational differences result from the C112R substitution in the N-terminal helix bundle and likely relate to a reduced ability of apoE4 to form tetramers, thereby increasing the concentration of functional apoE4 monomers, which gives rise to its higher lipid binding compared with apoE3.

ApoE2, ApoE3, and ApoE4 Differentially Stimulate APP Transcription and Aβ Secretion

Human apolipoprotein E (ApoE) apolipoprotein is primarily expressed in three isoforms (ApoE2, ApoE3, and ApoE4) that differ only by two residues. ApoE4 constitutes the most important genetic risk factor for Alzheimer's disease (AD), ApoE3 is neutral, and ApoE2 is protective. How ApoE isoforms influence AD pathogenesis, however, remains unclear. Using ES-cell-derived human neurons, we show that ApoE secreted by glia stimulates neuronal Aβ production with an ApoE4> ApoE3> ApoE2 potency rank order. We demonstrate that ApoE binding to ApoE receptors activates dual leucine-zipper kinase (DLK), a MAP-kinase kinase kinase that then activates MKK7 and ERK1/2 MAP kinases. Activated ERK1/2 induces cFos phosphorylation, stimulating the transcription factor AP-1, which in turn enhances transcription of amyloid-β precursor protein (APP) and thereby increases amyloid-β levels. This molecular mechanism also regulates APP transcription in mice invivo. Our data describe a novel signal transduction pathway in neurons whereby ApoE activates a non-canonical MAP kinase cascade that enhances APP transcription and amyloid-β synthesis.

Solution structure of discoidal high-density lipoprotein particles with a shortened apolipoprotein A-I.

High-density lipoprotein (HDL) particles are cholesterol and lipid transport containers. Mature HDL particles destined for the liver develop through the formation of intermediate discoidal HDL particles, which are the primary acceptors for cholesterol. Here we present the three-dimensional structure of reconstituted discoidal HDL (rdHDL) particles, using a shortened construct of human apolipoprotein A-I, determined from a combination of nuclear magnetic resonance (NMR), electron paramagnetic resonance (EPR) and transmission electron microscopy (TEM) data. The rdHDL particles feature a protein double belt surrounding a lipid bilayer patch in an antiparallel fashion. The integrity of this structure is maintained by up to 28 salt bridges and a zipper-like pattern of cation-π interactions between helices 4 and 6. To accommodate a hydrophobic interior, a gross 'right-to-right' rotation of the helices after lipidation is necessary. The structure reflects the complexity required for a shuttling container to hold a fluid lipid or cholesterol interior at a protein:lipid ratio of 1:50.

Altered cerebral insulin response in transgenic mice expressing the epsilon-4 allele of the human apolipoprotein E gene

Apolipoprotein E epsilon-4 (APOEε4 or APOE4), an allelic variation of the APOE gene, not only increases the risk of developing the late-onset form of Alzheimer’s disease (AD), but also influences the outcome of treatment. Indeed, data from clinical studies show that the beneficial effect of insulin on cognition is blunted in APOE4 carriers. To investigate how APOE impacts insulin response, we assessed the effects of an acute insulin injection in APOE3- and APOE4-targeted replacement mice that respectively express the human APOE3 or APOE4 isoform instead of the endogenous murine ApoE protein. We evaluated cognition, insulin signaling and proteins implicated in Aβ transport and tau phosphorylation in the cortex and brain capillaries. We found that a single acute insulin injection increased Akt pSer473 in APOE4 compared to APOE3 mice (+113% versus +78.5%), indicating that APOE4 carriage potentiates activation of insulin upstream signaling pathway in the brain. Insulin also led to decreased concentrations of the receptor for advanced glycation endproducts (RAGE) in brain capillaries in both groups of mice. Moreover, higher phosphorylation of tau at Ser202, one of the key markers of AD neuropathology, was observed in insulin-injected APOE4 mice (+44%), consistent with findings in human APOE4 carriers (+400% compared to non-carriers). Therefore, our data suggest that APOE4 carriage leads to an increased insulin-induced activation of cerebral Akt pathway, associated with higher AD-like tau neuropathology. Our results provide evidence of altered insulin signaling in APOE4 carriers as well as a possible mechanism to explain the absence of cognitive benefit from insulin therapy in these individuals.

Abstract 13218: Remodeling of High Density Lipoprotein Induced by CSL112: Structure and Function of the Remodeled Lipoproteins

Introduction: CSL112 is a novel formulation of human apolipoprotein A-I (apoA-I) suitable for intravenous infusion, currently in development for reducing the high rate of early recurrent events after acute myocardial infarction. Infusion of CSL112 into healthy volunteers causes HDL remodeling, with an immediate rise in lipid-poor apoA-I and a strong and immediate increase in the ABCA1-dependent cholesterol efflux capacity of plasma. Interaction of CSL112 with whole plasma or purified HDL gives rise to three new remodeled HDL species: large (L-HDL rem ) and small (S-HDL rem ) species, and lipid-poor apoA-I. Here we characterized the composition and morphology of remodeled HDL species and compared their ability to efflux cholesterol via different transporters. Methods and Results: CSL112 and HDL3, fluorescently labeled on protein or lipid, were incubated together for varying periods of time at 37°C. Resultant HDL particle size distribution was assessed by native gel electrophoresis followed by fluorescence imaging. Morphology of the individual purified products of HDL remodeling was examined by negative stain electron microscopy. We show that L-HDL rem is a large, spherical species composed of apoA-I and phospholipid from both native HDL and CSL112. S-HDL rem is a small, disc-shaped species composed of apoA-I from CSL112 that is smaller due to the loss of phospholipids. The smallest species, lipid-poor apoA-I, is composed of apoA-I from HDL and CSL112. Lipid-poor apoA-I and S-HDL rem were found to have the greatest ability to efflux cholesterol via ABCA1. While S-HDL rem and L-HDL rem showed comparable ability to efflux cholesterol via ABCG1, L-HDL rem mediated additional efflux via SR-B1 and by an aqueous diffusion process. Conclusion: The generation of these remodeled HDL species can account for the strong and immediate increase in cholesterol efflux capacity observed upon infusion of CSL112, and make CSL112 a promising approach for rapidly removing cholesterol from atherosclerotic plaque.

Structural changes induced by acidic pH in human apolipoprotein B-100.

Acidification in the endosome causes lipoprotein release by promoting a conformational change in the LDLR allowing its recycling and degradation of LDL. Notwithstanding conformational changes occurring in the LDLR have expanded considerably, structural changes occurring in LDL particles have not been fully explored yet. The objectives of the present work were to study structural changes occurring in apoB100 by infrared spectroscopy (IR) and also LDL size and morphology by dynamic light scattering (DLS) and electron microscopy (EM) at both pH 7.4 and 5.0. We determined by IR that pH acidification from 7.4 to 5.0, resembling that occurring within endosomal environment, induces a huge reversible structural rearrangement of apoB100 that is characterized by a reduction of beta-sheet content in favor of alpha-helix structures. Data obtained from DLS and EM showed no appreciable differences in size and morphology of LDL. These structural changes observed in apoB100, which are likely implied in particle release from lipoprotein receptor, also compromise the apoprotein stability what would facilitate LDL degradation. In conclusion, the obtained results reveal a more dynamic picture of the LDL/LDLR dissociation process than previously perceived and provide new structural insights into LDL/LDLR interactions than can occur at endosomal low-pH milieu.

APOBEC3 deletion increases the risk of breast cancer: a meta-analysis.

Recently, a deletion in the human apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like 3 (APOBEC3) gene cluster has been associated with a modest increased risk of breast cancer, but studies yielded inconsistent results. Therefore we performed a meta-analysis to derive a more precise conclusion. Six studies including 18241 subjects were identified by searching PubMed and Embase databases from inception to April 2016. Pooled odds ratios (ORs) and corresponding 95% confidence intervals (CIs) were evaluated under allele contrast, dominant, recessive, homozygous, and heterozygous models. All the analyses suggested a correlation of APOBEC3 deletion with increased breast cancer risk (D vs I: OR = 1.29, 95% CI = 1.23-1.36; D/D+I/D vs I/I: OR = 1.34, 95% CI = 1.26-1.43; D/D vs I/D+ I/I: OR = 1.51, 95% CI = 1.36-1.68; D/D vs I/I: OR = 1.75, 95% CI= 1.56-1.95; I/D vs I/I: OR = 1.28, 95% CI = 1.19-1.36). Stratified analysis by ethnicity showed that the relationship is stronger and more stable in Asians. In summary, our current work indicated that APOBEC3 copy number variations might have a good screening accuracy for breast cancer.

Enhanced HDL Functionality in Small HDL Species Produced Upon Remodeling of HDL by Reconstituted HDL, CSL112

Rationale: CSL112, human apolipoprotein A-I (apoA-I) reconstituted with phosphatidylcholine, is known to cause a dramatic rise in small high-density lipoprotein (HDL). Objective: To explore the mechanisms by which the formation of small HDL particles is induced by CSL112. Methods and Results: Infusion of CSL112 into humans caused elevation of 2 small diameter HDL fractions and 1 large diameter fraction. Ex vivo studies showed that this remodeling does not depend on lipid transfer proteins or lipases. Rather, interaction of CSL112 with purified HDL spontaneously gave rise to 3 HDL species: a large, spherical species composed of apoA-I from native HDL and CSL112; a small, disc-shaped species composed of apoA-I from CSL112, but smaller because of the loss of phospholipids; and the smallest species, lipid-poor apoA-I composed of apoA-I from HDL and CSL112. Time-course studies suggest that remodeling occurs by an initial fusion of CSL112 with HDL and subsequent fission leading to the smaller forms. Functional studies showed that ATP-binding cassette transporter 1–dependent cholesterol efflux and anti-inflammatory effects in whole blood were carried by the 2 small species with little activity in the large species. In contrast, the ability to inactivate lipid hydroperoxides in oxidized low-density lipoprotein was carried predominantly by the 2 largest species and was low in lipid-poor apoA-I. Conclusions: We have described a mechanism for the formation of small, highly functional HDL species involving spontaneous fusion of discoidal HDL with spherical HDL and subsequent fission. Similar remodeling is likely to occur during the life cycle of apoA-I in vivo.

Structural changes induced by acidic pH in human apolipoprotein B-100

Structural changes induced by acidic pH in human apolipoprotein B-100

PH- and surface pressure-dependant adsorption of human apolipoprotein A1 at solid/liquid and gas/liquid interfaces

pH- and surface pressure-dependant adsorption of human apolipoprotein A1 at solid/liquid and gas/liquid interfaces

Conservation in the phylum of the local homology of apolipoproteins with the thyroid hormone plasma carriers.

Thyroxine-binding globulin (TBG), transthyretin (TTR), albumin (HSA), plus other plasmatic proteins, which include apolipoproteins, can bind and transport thyroid hormones (TH). In 1994, a 5-residue motif (Y, L/I/M, X, X, V/L/I) conserved in human TBG, TTR, HSA, and human and animal apolipoproteins was identified. Recently, we noticed that a number of residues upstream and downstream that motif are also conserved.We tested in silico the conservation of this larger motif in the many additional animal sequences of TH plasma carriers discovered after 1994. To this aim, we searched for the occurrence of the "new" motif in human and animal apolipoprotein and non-apolipoprotein TH-binding plasmatic proteins, and in a group of randomly selected proteins (2918 sequences from 56 species) not known as TH binders.Our results confirm the conservation of the "new" motif, associated with TH binding, in a total of 426 sequences analyzed (220 belonging to 169 apolipoproteins from 69 species, 206 belonging to 123 nonapolipoproteins from 54 species). Additionally, we found that within such conserved segments some differences between groups of TH plasma carriers exist. Interestingly, number and type of differences appear related to the affinity of each carrier for thyroid hormones. No occurrence of the motif was found in control proteins (alpha- and beta-tubulin, eosinophil cationic protein, endothelin-1, -2 and -3, IgG receptor, tropomyosin, Wnt inhibitory factor 1, erythropoietin, insulin and haptoglobin).Maintenance of a TH-binding domain in apolipoproteins throughout the phylum should be not less important that maintenance of the lipid binding domain.