scholarly journals PRAP1 is a novel lipid binding protein that promotes lipid absorption by facilitating MTTP-mediated lipid transport

2020 ◽  
pp. jbc.RA120.015002 ◽  
Author(s):  
Hubert Peng ◽  
Tzu-Yuan Chiu ◽  
Yu-Jen Liang ◽  
Chia-Jen Lee ◽  
Chih-Syuan Liu ◽  
...  
Keyword(s):  
Ternary Complex ◽  
Microsomal Triglyceride Transfer Protein ◽  
Lipid Binding ◽  
Lipid Absorption ◽  
Lipid Transport ◽  
Single Amino Acid ◽  
Chow Diet ◽  
Lipid Transfer ◽  
Plasma Fractions ◽  
Fat Diets

Microsomal triglyceride transfer protein (MTTP) is an endoplasmic reticulum (ER) resident protein that is essential for the assembly and secretion of triglyceride (TG)-rich, apoB-containing lipoproteins. Although the function and structure of mammalian MTTP have been extensively studied, how exactly MTTP transfers lipids to lipid acceptors and whether there are other biomolecules involved in MTTP-mediated lipid transport remain elusive. Here we identify a role in this process for the poorly characterized protein PRAP1. We report that PRAP1 and MTTP are partially co-localized in the ER. We observe that PRAP1 directly binds to TG and facilitates MTTP-mediated lipid transfer. A single amino acid mutation at position 85 (E85V) impairs PRAP1's ability to form a ternary complex with TG and MTTP, as well as impairs its ability to facilitate MTTP-mediated apoB-containing lipoprotein assembly and secretion, suggesting that the ternary complex formation is required for PRAP1 to facilitate MTTP-mediated lipid transport. PRAP1 is detectable in chylomicron/VLDL-rich plasma fractions, suggesting that MTTP recognizes PRAP1-bound TG as a cargo and transfers TG along with PRAP1 to lipid acceptors. Both PRAP1 deficient and the E85V knock-in mutant mice fed a chow diet manifested an increase in the length of their small intestines, likely to compensate for challenges in absorbing lipid. Interestingly, both genetically modified mice gained significantly less body weight and fat mass when on high fat diets compared to littermate controls and were prevented from hepatosteatosis. Together, this study provides evidence that PRAP1 plays an important role in MTTP-mediated lipid transport and lipid absorption.

Lipoprotein assembly and function in an evolutionary perspective

2010 ◽  
Vol 1 (2) ◽  
pp. 165-183 ◽  
Author(s):  
Dick J. Van der Horst ◽  
Kees W. Rodenburg
Keyword(s):  
Lipid Transfer Protein ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Microsomal Triglyceride Transfer Protein ◽  
Lipid Binding ◽  
Lipid Transfer ◽  
Transfer Protein ◽  
Density Lipoprotein Receptor ◽  
Exchangeable Basis ◽  
And Function

AbstractCirculatory fat transport in animals relies on members of the large lipid transfer protein (LLTP) superfamily, including mammalian apolipoprotein B (apoB) and insect apolipophorin II/I (apoLp-II/I). ApoB and apoLp-II/I, constituting the structural (non-exchangeable) basis for the assembly of various lipoproteins, acquire lipids through microsomal triglyceride-transfer protein, another LLTP family member, and bind them by means of amphipathic α-helical and β-sheet structural motifs. Comparative research reveals that LLTPs evolved from the earliest animals and highlights the structural adaptations in these lipid-binding proteins. Thus, in contrast to apoB, apoLp-II/I is cleaved post-translationally by a furin, resulting in the appearance of two non-exchangeable apolipoproteins in the single circulatory lipoprotein in insects, high-density lipophorin (HDLp). The remarkable structural similarities between mammalian and insect lipoproteins notwithstanding important functional differences relate to the mechanism of lipid delivery. Whereas in mammals, partial delipidation of apoB-containing lipoproteins eventually results in endocytic uptake of their remnants, mediated by members of the low-density lipoprotein receptor (LDLR) family, and degradation in lysosomes, insect HDLp functions as a reusable lipid shuttle capable of alternate unloading and reloading of lipid. Also, during muscular efforts (flight activity), an HDLp-based lipoprotein shuttle provides for the transport of lipid for energy generation. Although a lipophorin receptor – a homolog of LDLR – was identified that mediates endocytic uptake of HDLp during specific developmental periods, the endocytosed lipoprotein appears to be recycled in a transferrin-like manner. These data highlight that the functional adaptations in the lipoprotein lipid carriers in mammals and insects also emerge with regard to the functioning of their cognate receptors.

scholarly journals Regulation of microsomal triglyceride transfer protein by apolipoprotein A-IV in newborn swine intestinal epithelial cells

2011 ◽  
Vol 300 (2) ◽  
pp. G357-G363 ◽  
Author(s):  
Ying Yao ◽  
Song Lu ◽  
Yue Huang ◽  
Casey C. Beeman-Black ◽  
Rena Lu ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Intestinal Epithelial Cells ◽  
Microsomal Triglyceride Transfer Protein ◽  
Lipid Absorption ◽  
Intestinal Epithelial ◽  
Lipid Transfer ◽  
Protein Levels ◽  
Transfer Protein ◽  
Transfer Activity ◽  
The Impact

Apolipoprotein (apo) A-IV overexpression enhances chylomicron (CM) assembly and secretion in newborn swine intestinal epithelial cells by producing larger particles (Lu S, Yao Y, Cheng X, Mitchell S, Leng S, Meng S, Gallagher JW, Shelness GS, Morris GS, Mahan J, Frase S, Mansbach CM, Weinberg RB, Black DD. J Biol Chem 281: 3473–3483, 2006). To determine the impact of apo A-IV on microsomal triglyceride transfer protein (MTTP), IPEC-1 cell lines containing a tetracycline-regulatable expression system were used to overexpress native swine apo A-IV and “piglike” human apo A-IV, a mutant human apo A-IV with deletion of the EQQQ-rich COOH-terminus, previously shown to upregulate basolateral triglyceride (TG) secretion 5-fold and 25-fold, respectively. Cells were incubated 24 h with and without doxycycline and oleic acid (OA, 0.8 mM). Overexpression of the native swine apo A-IV and piglike human apo A-IV increased MTTP lipid transfer activity by 39.7% ( P = 0.006) and 53.6% ( P = 0.0001), respectively, compared with controls. Changes in mRNA and protein levels generally paralleled changes in activity. Interestingly, native swine apo A-IV overexpression also increased MTTP large subunit mRNA, protein levels, and lipid transfer activity in the absence of OA, suggesting a mechanism not mediated by lipid absorption. Overexpression of piglike human apo A-IV significantly increased partitioning of radiolabeled OA from endoplasmic reticulum (ER) membrane to lumen, suggesting increased net transfer of membrane TG to luminal particles. These results suggest that the increased packaging of TG into nascent CMs in the ER lumen, induced by apo A-IV, is associated with upregulation of MTTP activity at the pretranslational level. Thus MTTP is regulated by apo A-IV in a manner to promote increased packaging of TG into the CM core, which may be important in neonatal fat absorption.

scholarly journals Why does the gut choose apolipoprotein B48 but not B100 for chylomicron formation?

2008 ◽  
Vol 294 (1) ◽  
pp. G344-G352 ◽  
Author(s):  
Chun-Min Lo ◽  
Brian K. Nordskog ◽  
Andromeda M. Nauli ◽  
Shuqin Zheng ◽  
Sarah B. vonLehmden ◽  
...  
Keyword(s):  
Microsomal Triglyceride Transfer Protein ◽  
Lipid Absorption ◽  
Lipid Transfer ◽  
Wild Type ◽  
Very Low Density Lipoproteins ◽  
Apo B ◽  
Transfer Protein ◽  
Apolipoprotein B48 ◽  
Lipid Dose ◽  
Chylomicron Formation

Chylomicrons produced by the human gut contain apolipoprotein (apo) B48, whereas very-low-density lipoproteins made by the liver contain apo B100. To study how these molecules function during lipid absorption, we examined the process as it occurs in apobec-1 knockout mice (able to produce only apo B100; KO) and in wild-type mice (of which the normally functioning intestine makes apo B48, WT). Using the lymph fistula model, we studied the process of lipid absorption when animals were intraduodenally infused with a lipid emulsion (4 or 6 μmol/h of triolein). KO mice transported triacylglycerol (TG) as efficiently as WT mice when infused with the lower lipid dose; when infused with 6 μmol/h of triolein, however, KO mice transported significantly less TG to lymph than WT mice, leading to the accumulation of mucosal TG. Interestingly, the size of lipoprotein particles from both KO and WT mice were enlarged to chylomicron-size particles during absorption of the higher dose. These increased-size particles produced by KO mice were not associated with increased apo AIV secretion. However, we found that the gut of the KO mice secreted fewer apo B molecules to lymph (compared with WT), during both fasting and lipid infusion, leading us to conclude that the KO gut produced fewer numbers of TG-rich lipoproteins (including chylomicron) than the wild-type animals. The reduced apo B secretion in KO mice was not related to reduced microsomal triglyceride transfer protein lipid transfer activity. We propose that apo B48 is the preferred protein for the gut to coat chylomicrons to ensure efficient chylomicron formation and lipid absorption.

scholarly journals The crystal structure of human microsomal triglyceride transfer protein

2019 ◽  
Vol 116 (35) ◽  
pp. 17251-17260 ◽  
Author(s):  
Ekaterina I. Biterova ◽  
Michail N. Isupov ◽  
Ronan M. Keegan ◽  
Andrey A. Lebedev ◽  
Anil A. Sohail ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Structural Information ◽  
Neutral Lipids ◽  
Microsomal Triglyceride Transfer Protein ◽  
Lipid Binding ◽  
Rational Drug Design ◽  
Lipid Transfer ◽  
Very Low Density Lipoproteins ◽  
Transfer Protein ◽  
Binding Cavity

Microsomal triglyceride transfer protein (MTP) plays an essential role in lipid metabolism, especially in the biogenesis of very low-density lipoproteins and chylomicrons via the transfer of neutral lipids and the assembly of apoB-containing lipoproteins. Our understanding of the molecular mechanisms of MTP has been hindered by a lack of structural information of this heterodimeric complex comprising an MTPα subunit and a protein disulfide isomerase (PDI) β-subunit. The structure of MTP presented here gives important insights into the potential mechanisms of action of this essential lipid transfer molecule, structure-based rationale for previously reported disease-causing mutations, and a means for rational drug design against cardiovascular disease and obesity. In contrast to the previously reported structure of lipovitellin, which has a funnel-like lipid-binding cavity, the lipid-binding site is encompassed in a β-sandwich formed by 2 β-sheets from the C-terminal domain of MTPα. The lipid-binding cavity of MTPα is large enough to accommodate a single lipid. PDI independently has a major role in oxidative protein folding in the endoplasmic reticulum. Comparison of the mechanism of MTPα binding by PDI with previously published structures gives insights into large protein substrate binding by PDI and suggests that the previous structures of human PDI represent the “substrate-bound” and “free” states rather than differences arising from redox state.

scholarly journals The lipid transfer protein Saposin B does not directly bind CD1d for lipid antigen loading

2019 ◽  
Vol 4 ◽  
pp. 117
Author(s):  
Maria Shamin ◽  
Tomasz H. Benedyk ◽  
Stephen C. Graham ◽  
Janet E. Deane
Keyword(s):  
Lipid Transfer Protein ◽  
Direct Interaction ◽  
Lipid Binding ◽  
Lipid Transfer ◽  
Binding Assays ◽  
Lipid Transfer Proteins ◽  
Lipid Antigens ◽  
Saposin B ◽  
Protein Components ◽  
Lipid Loading

Background: Lipid antigens are presented on the surface of cells by the CD1 family of glycoproteins, which have structural and functional similarity to MHC class I molecules. The hydrophobic lipid antigens are embedded in membranes and inaccessible to the lumenal lipid-binding domain of CD1 molecules. Therefore, CD1 molecules require lipid transfer proteins for lipid loading and editing. CD1d is loaded with lipids in late endocytic compartments, and lipid transfer proteins of the saposin family have been shown to play a crucial role in this process. However, the mechanism by which saposins facilitate lipid binding to CD1 molecules is not known and is thought to involve transient interactions between protein components to ensure CD1-lipid complexes can be efficiently trafficked to the plasma membrane for antigen presentation. Of the four saposin proteins, the importance of Saposin B (SapB) for loading of CD1d is the most well-characterised. However, a direct interaction between CD1d and SapB has yet to be described. Methods: In order to determine how SapB might load lipids onto CD1d, we used purified, recombinant CD1d and SapB and carried out a series of highly sensitive binding assays to monitor direct interactions. We performed equilibrium binding analysis, chemical cross-linking and co-crystallisation experiments, under a range of different conditions. Results: We could not demonstrate a direct interaction between SapB and CD1d using any of these binding assays. Conclusions: This work establishes comprehensively that the role of SapB in lipid loading does not involve direct binding to CD1d. We discuss the implication of this for our understanding of lipid loading of CD1d and propose several factors that may influence this process.

scholarly journals The structure and flexibility analysis of the Arabidopsis Synaptotagmin 1 reveal the basis of its regulation at membrane contact sites

2021 ◽  
Author(s):  
Juan Luis Benavente ◽  
Dritan Siliqi ◽  
Lourdes Infantes ◽  
Laura Lagartera ◽  
Alberto Mills ◽  
...  
Keyword(s):  
Cell Function ◽  
Lipid Extraction ◽  
Membrane Lipid ◽  
Lipid Binding ◽  
Lipid Transfer ◽  
C2 Domains ◽  
Membrane Contact Sites ◽  
Contact Sites ◽  
Cellular Environment ◽  
Synaptotagmin 1

Cell function requires the maintenance of membrane lipid homeostasis as changes in cellular environment unbalance this equilibrium. The non-vesicular lipid transfer at endoplasmic reticulum (ER) and plasma membrane (PM) contact sites (CS) is central to restore it. Extended synaptotagmins (E-Syts) are ER proteins that play a central role in this process as they act as molecular tethers with PM and as lipid transfer proteins between these organelles. E-Syts are constitutively anchored to the ER through an N-terminal hydrophobic segment and bind to the PM via C-terminal C2 domains. In plants, synaptotagmins (SYTs) are orthologous of E-Syts and regulate the ER-PM communication by the activity of their two C2 domains in response to abiotic stresses. We have combined macromolecular crystallography, small-angle X-ray scattering, structural bioinformatics and biochemical data to analyze the regulation of plant synaptotagmin 1 (SYT1). Our data show that the binding of SYT1 to the PM is regulated by the interaction of the first C2 domain through a Ca2+-dependent lipid binding site and by a site for phosphorylated forms of phosphatidylinositol in such a way that two different molecular signals are integrated in response to stress. In addition, our data show that SYT1 is highly flexible by virtue of up to three hinge points, including one that connects the two C2 domains. This feature provides conformational freedom to SYT1 to define a large and complementary interaction surface with the PM. This structural plasticity, in turn, may facilitate lipid extraction, protein loading and subsequent transfer between PM and ER.

scholarly journals Redox proteomics of stomatal immunity reveals role of a lipid transfer protein in plant defense

2020 ◽  
Author(s):  
Kelly M Balmant ◽  
Sheldon R Lawrence ◽  
Benjamin V Duong ◽  
Fanzhao Zhu ◽  
Ning Zhu ◽  
...  
Keyword(s):  
Plant Defense ◽  
Lipid Transfer Protein ◽  
Guard Cells ◽  
Lipid Binding ◽  
Lipid Transfer ◽  
Redox Proteomics ◽  
Transfer Protein ◽  
Redox Responsive ◽  
Redox Sensors ◽  
Thiol Redox

ABSTRACTRedox-based post-translational modifications (PTMs) involving protein cysteine residues as redox sensors are important to various physiological processes. However, little is known about redox-sensitive proteins in guard cells and their functions in stomatal immunity. In this study, we applied an integrative protein labeling method cysTMTRAQ and identified guard cell proteins that were altered by thiol redox PTMs in response to a bacterial flagellin peptide flg22. In total, eight, seven and 20 potential redox-responsive proteins were identified in guard cells treated with flg22 for 15, 30 and 60 min, respectively. The proteins fall into several functional groups including photosynthesis, lipid binding, oxidation-reduction, and defense. Among the proteins, a lipid transfer protein (LTP)-II was confirmed to be redox-responsive and involved in plant resistance to Pseudomonas syringe pv. tomato DC3000. This study not only creates an inventory of potential redox-sensitive proteins in flg22 signal transduction in guard cells, but also highlights the relevance of the lipid transfer protein in plant defense against the bacterial pathogens.Sentence summaryThiol-redox proteomics identified potential redox sensors important in stomatal immunity, and a lipid transfer protein was characterized to function as a redox sensor in plant immune response.

scholarly journals Black, pinto and white beans lower hepatic lipids in hamsters fed high fat diets by excretion of bile acids

2020 ◽  
Vol 2 (1) ◽  
Author(s):  
Priscila L. S. Alves ◽  
Jose De J Berrios ◽  
James Pan ◽  
Wallace H. Yokoyama
Keyword(s):  
Lithocholic Acid ◽  
Plasma Cholesterol ◽  
Liver Cholesterol ◽  
Control Group ◽  
Chow Diet ◽  
High Fat ◽  
Bean Flour ◽  
Freeze Dried ◽  
High Fat Diets ◽  
Fat Diets

Abstract Hypolipidemic and anti-obesity properties of extruded black, pinto and white beans were examined in male Syrian hamsters fed high fat diets for 3 weeks. The efficiency and effectiveness of extruder processing to eliminate heat-labile antinutrients were also determined. Hamsters fed a high fat diet containing 40% extruded black beans gained the same amount of weight as animals on a low-fat chow diet (based on AIN-93G). Total plasma cholesterol of the hamsters fed bean flour- based diets tended to be lower, ranging between 298 and 356 mg/dL, compared to 365 mg/dL for the control group and total and free liver cholesterol was about 50 and 33% lower, respectively, in the livers (freeze-dried) of hamsters fed the black turtle bean (BB) and pinto bean (PB) diets. The nitrogen content of the feces from hamsters fed the PB and white bean (WB) diets were 2.6x higher than the control and the excretion of deoxycholic acid (DCA) and lithocholic acid (LCA) was higher in all legume fed hamsters The results show that extrusion efficiently and economically reduces anti-nutritive factors that inhibit the digestion and absorption of proteins and carbohydrates. Graphical abstract

scholarly journals Structural characterization and in vitro lipid binding studies of non-specific lipid transfer protein 1 (nsLTP1) from fennel (Foeniculum vulgare) seeds

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Mekdes Megeressa ◽  
Bushra Siraj ◽  
Shamshad Zarina ◽  
Aftab Ahmed
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
3D Structure ◽  
Lipid Binding ◽  
Lipid Transfer ◽  
Binding Studies ◽  
Foeniculum Vulgare ◽  
Complete Amino Acid Sequence ◽  
Specific Lipid

AbstractNon-specific lipid transfer proteins (nsLTPs) are cationic proteins involved in intracellular lipid shuttling in growth and reproduction, as well as in defense against pathogenic microbes. Even though the primary and spatial structures of some nsLTPs from different plants indicate their similar features, they exhibit distinct lipid-binding specificities signifying their various biological roles that dictate further structural study. The present study determined the complete amino acid sequence, in silico 3D structure modeling, and the antiproliferative activity of nsLTP1 from fennel (Foeniculum vulgare) seeds. Fennel is a member of the family Umbelliferae (Apiaceae) native to southern Europe and the Mediterranean region. It is used as a spice medicine and fresh vegetable. Fennel nsLTP1 was purified using the combination of gel filtration and reverse-phase high-performance liquid chromatography (RP-HPLC). Its homogeneity was determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry. The purified nsLTP1 was treated with 4-vinyl pyridine, and the modified protein was then digested with trypsin. The complete amino acid sequence of nsLTP1 established by intact protein sequence up to 28 residues, overlapping tryptic peptides, and cyanogen bromide (CNBr) peptides. Hence, it is confirmed that fennel nsLTP1 is a 9433 Da single polypeptide chain consisting of 91 amino acids with eight conserved cysteines. Moreover, the 3D structure is predicted to have four α-helices interlinked by three loops and a long C-terminal tail. The lipid-binding property of fennel nsLTP1 is examined in vitro using fluorescent 2-p-toluidinonaphthalene-6-sulfonate (TNS) and validated using a molecular docking study with AutoDock Vina. Both of the binding studies confirmed the order of binding efficiency among the four studied fatty acids linoleic acid > linolenic acid > Stearic acid > Palmitic acid. A preliminary screening of fennel nsLTP1 suppressed the growth of MCF-7 human breast cancer cells in a dose-dependent manner with an IC50 value of 6.98 µM after 48 h treatment.

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