共查询到20条相似文献,搜索用时 31 毫秒
1.
Ben-Zeev O Hosseini M Lai CM Ehrhardt N Wong H Cefalù AB Noto D Averna MR Doolittle MH Péterfy M 《Journal of lipid research》2011,52(6):1162-1169
Lipase maturation factor 1 (Lmf1) is an endoplasmic reticulum (ER) membrane protein involved in the posttranslational folding and/or assembly of lipoprotein lipase (LPL) and hepatic lipase (HL) into active enzymes. Mutations in Lmf1 are associated with diminished LPL and HL activities ("combined lipase deficiency") and result in severe hypertriglyceridemia in mice as well as in human subjects. Here, we investigate whether endothelial lipase (EL) also requires Lmf1 to attain enzymatic activity. We demonstrate that cells harboring a (cld) loss-of-function mutation in the Lmf1 gene are unable to generate active EL, but they regain this capacity after reconstitution with the Lmf1 wild type. Furthermore, we show that cellular EL copurifies with Lmf1, indicating their physical interaction in the ER. Finally, we determined that post-heparin phospholipase activity in a patient with the LMF1(W464X) mutation is reduced by more than 95% compared with that in controls. Thus, our study indicates that EL is critically dependent on Lmf1 for its maturation in the ER and demonstrates that Lmf1 is a required factor for all three vascular lipases, LPL, HL, and EL. 相似文献
2.
Over a third of the US adult population has hypertriglyceridemia, resulting in an increased risk of atherosclerosis, pancreatitis, and metabolic syndrome. Lipoprotein lipase (LPL), a dimeric enzyme, is the main lipase responsible for TG clearance from the blood after food intake. LPL requires an endoplasmic reticulum (ER)-resident, transmembrane protein known as lipase maturation factor 1 (LMF1) for secretion and enzymatic activity. LMF1 is believed to act as a client specific chaperone for dimeric lipases, but the precise mechanism by which LMF1 functions is not understood. Here, we examine which domains of LMF1 contribute to dimeric lipase maturation by assessing the function of truncation variants. N-terminal truncations of LMF1 show that all the domains are necessary for LPL maturation. Fluorescence microscopy and protease protection assays confirmed that these variants were properly oriented in the ER. We measured cellular levels of LMF1 and found that it is expressed at low levels and each molecule of LMF1 promotes the maturation of 50 or more molecules of LPL. Thus we provide evidence for the critical role of the N-terminus of LMF1 for the maturation of LPL and relevant ratio of chaperone to substrate. 相似文献
3.
4.
Li S Nagothu K Ranganathan G Ali SM Shank B Gokden N Ayyadevara S Megyesi J Olivecrona G Chugh SS Kersten S Portilla D 《American journal of physiology. Renal physiology》2012,303(3):F437-F448
Peroxisome proliferator-activated receptor-α (PPARα) activation attenuates cisplatin (CP)-mediated acute kidney injury by increasing fatty acid oxidation, but mechanisms leading to reduced renal triglyceride (TG) accumulation could also contribute. Here, we investigated the effects of PPARα and CP on expression and enzyme activity of kidney lipoprotein lipase (LPL) as well as on expression of angiopoietin protein-like 4 (Angptl4), glycosylphosphatidylinositol-anchored-HDL-binding protein (GPIHBP1), and lipase maturation factor 1 (Lmf1), which are recognized as important proteins that modulate LPL activity. CP caused a 40% reduction in epididymal white adipose tissue (WAT) mass, with a reduction of LPL expression and activity. CP also reduced kidney LPL expression and activity. Angptl4 mRNA levels were increased by ninefold in liver and kidney tissue and by twofold in adipose tissue of CP-treated mice. Western blots of two-dimensional gel electrophoresis identified increased expression of a neutral pI Angptl4 protein in kidney tissue of CP-treated mice. Immunolocalization studies showed reduced staining of LPL and increased staining of Angptl4 primarily in proximal tubules of CP-treated mice. CP also increased TG accumulation in kidney tissue, which was ameliorated by PPARα ligand. In summary, a PPARα ligand ameliorates CP-mediated nephrotoxicity by increasing LPL activity via increased expression of GPHBP1 and Lmf1 and by reducing expression of Angptl4 protein in the proximal tubule. 相似文献
5.
Lipoprotein lipase (LPL) is a secreted lipase that clears triglycerides from the blood. Proper LPL folding and exit from the endoplasmic reticulum (ER) require lipase maturation factor 1 (LMF1), an ER‐resident transmembrane protein, but the mechanism involved is unknown. We used proteomics to identify LMF1‐binding partners necessary for LPL secretion in HEK293 cells and found these to include oxidoreductases and lectin chaperones, suggesting that LMF1 facilitates the formation of LPL's five disulfide bonds. In accordance with this role, we found that LPL aggregates in LMF1‐deficient cells due to the formation of incorrect intermolecular disulfide bonds. Cells lacking LMF1 were hypersensitive to depletion of glutathione, but not DTT treatment, suggesting that LMF1 helps reduce the ER. Accordingly, we found that loss of LMF1 results in a more oxidized ER. Our data show that LMF1 has a broader role than simply folding lipases, and we identified fibronectin and the low‐density lipoprotein receptor (LDLR) as novel LMF1 clients that contain multiple, non‐sequential disulfide bonds. We conclude that LMF1 is needed for secretion of some ER client proteins that require reduction of non‐native disulfides during their folding. 相似文献
6.
Combined lipase deficiency (cld) is a recessive, lethal mutation specific to the t
w73
haplotype on mouse Chromosome 17. While the cld mutation results in lipase proteins that are inactive, aggregated, and retained in the endoplasmic reticulum (ER), it maps
separately from the lipase structural genes. We have narrowed the gene critical region by about 50% using the t
w18
haplotype for deletion mapping and a recombinant chromosome used originally to map cld with respect to the phenotypic marker tf. The region now extends from 22 to 25.6 Mbp on the wild-type chromosome, currently containing 149 genes and 50 expressed sequence
tags (ESTs). To identify the affected gene, we have selected candidates based on their known role in associated biological
processes, cellular components, and molecular functions that best fit with the predicted function of the cld gene. A secondary approach was based on differences in mRNA levels between mutant (cld/cld) and unaffected (+/cld) cells. Using both approaches, we have identified seven functional candidates with an ER localization and/or an involvement
in protein maturation and folding that could explain the lipase deficiency, and six expression candidates that exhibit large
differences in mRNA levels between mutant and unaffected cells. Significantly, two genes were found to be candidates with
regard to both function and expression, thus emerging as the strongest candidates for cld. We discuss the implications of our mapping results and our selection of candidates with respect to other genes, deletions,
and mutations occurring in the cld critical region. 相似文献
7.
Lindegaard ML Olivecrona G Christoffersen C Kratky D Hannibal J Petersen BL Zechner R Damm P Nielsen LB 《Journal of lipid research》2005,46(11):2339-2346
Placenta expresses various lipase activities. However, a detailed characterization of the involved genes and proteins is lacking. In this study, we compared the expression of endothelial lipase (EL) and LPL in human term placenta. When placental protein extracts were separated by heparin-Sepharose affinity chromatography, the EL protein eluted as a single peak without detectable phospholipid or triglyceride (TG) lipase activity. The major portion of LPL protein eluted slightly after EL. This peak also had no lipase activity and most likely contained monomeric LPL. Fractions eluting at a higher NaCl concentration contained small amounts of LPL protein (most likely dimeric LPL) and had substantial TG lipase activity. In situ hybridization studies showed EL mRNA expression in syncytiotrophoblasts and endothelial cells and LPL mRNA in syncytiotrophoblasts. In contrast, immunohistochemistry showed EL and LPL protein associated with both cell types. In mouse placentas, lack of LPL expression resulted in increased EL mRNA expression. These results suggest that the cellular expression of EL and LPL in human placenta is different. Nevertheless, the two lipases might have overlapping functions in the mouse placenta. Our data also suggest that the major portions of both proteins are stored in an inactive form in human term placenta. 相似文献
8.
Mark H. Doolittle Saskia B. Neher Osnat Ben-Zeev Jo Ling-liao Ciara M. Gallagher Maryam Hosseini Fen Yin Howard Wong Peter Walter Mikl��s P��terfy 《The Journal of biological chemistry》2009,284(48):33623-33633
Lipase maturation factor 1 (LMF1) is predicted to be a polytopic protein localized to the endoplasmic reticulum (ER) membrane. It functions in the post-translational attainment of enzyme activity for both lipoprotein lipase and hepatic lipase. By using transmembrane prediction methods in mouse and human orthologs, models of LMF1 topology were constructed and tested experimentally. Employing a tagging strategy that used insertion of ectopic glycan attachment sites and terminal fusions of green fluorescent protein, we established a five-transmembrane model, thus dividing LMF1 into six domains. Three domains were found to face the cytoplasm (the amino-terminal domain and loops B and D), and the other half was oriented to the ER lumen (loops A and C and the carboxyl-terminal domain). This representative model shows the arrangement of an evolutionarily conserved domain within LMF1 (DUF1222) that is essential to lipase maturation. DUF1222 comprises four of the six domains, with the two largest ones facing the ER lumen. We showed for the first time, using several naturally occurring variants featuring DUF1222 truncations, that Lmf1 interacts physically with lipoprotein lipase and hepatic lipase and localizes the lipase interaction site to loop C within DUF1222. We discuss the implication of our results with regard to lipase maturation and DUF1222 domain structure. 相似文献
9.
Garenc C Aubert S Laroche J Bergeron J Gagné C Rousseau F Julien P 《Biochemical and biophysical research communications》2006,344(2):588-596
In Eastern Québec, two major lipoprotein lipase (LPL) gene mutations, P207L and G188E, lead to complete LPL deficiency in homozygote subjects and contribute to elevated predisposition to hypertriglyceridemia in heterozygotes. First, we determined the allele frequencies of LPL (D9N, G188E, P207L, D250N, N291S, and S447X), APOE (C112R and C158R), PPARalpha (L162V), and PPARgamma2 (P12A) single nucleotide polymorphisms (SNPs) in a random-based cohort of the metropolitan Québec city area. Second, we compared the LPL X447 allele frequencies observed in the random cohort and in a cohort of LPL P207L deficient patients. In the random cohort, the LPL N9 rare allele exhibited a higher prevalence than previously expected (p=0.0001). The LPL X447 allele frequency was lower in the patient cohort (Freq: 4.4%) than in the random cohort (Freq: 11.2%) (p=0.0001). These results reveal the importance of genetic screening for LPL gene mutations D9N and S447X in a population at risk to develop hypertriglyceridemia. 相似文献
10.
Griffon N Budreck EC Long CJ Broedl UC Marchadier DH Glick JM Rader DJ 《Journal of lipid research》2006,47(8):1803-1811
The triglyceride (TG) lipase gene subfamily, consisting of LPL, HL, and endothelial lipase (EL), plays a central role in plasma lipoprotein metabolism. Compared with LPL and HL, EL is relatively more active as a phospholipase than as a TG lipase. The amino acid loop or "lid" covering the catalytic site has been implicated as the basis for the difference in substrate specificity between HL and LPL. To determine the role of the lid in the substrate specificity of EL, we studied EL in comparison with LPL by mutating specific residues of the EL lid and exchanging their lids. Mutation studies showed that amphipathic properties of the lid contribute to substrate specificity. Exchanging lids between LPL and EL only partially shifted the substrate specificity of the enzymes. Studies of a double chimera possessing both the lid and the C-terminal domain (C-domain) of EL in the LPL backbone showed that the role of the lid in determining substrate specificity does not depend on the nature of the C-domain of the lipase. Using a kinetic assay, we showed an additive effect of the EL lid on the apparent affinity for HDL(3) in the presence of the EL C-domain. 相似文献
11.
Rats subcutaneously implanted with AH109A hepatoma cells show hyperlipidemia with high concentrations of serum triglyceride
and nonesterified fatty acid, suppression of lipoprotein lipase (LPL), and elevation of hormone-sensitive lipase (HSL) activities
during the growth of the hepatoma. Supplementation of the diet with sulfur amino acids such as l-methionine (Met) and l-cystine (Cys) improved hyperlipidemia by restoring LPL and HSL activities. In the present study, we have attempted to examine
the effects of sulfur amino acids on the activity and mRNA level of LPL and the activity of HSL using 3T3-L1 cells, which
are known to differentiate to adipocytes. The adipocytes were incubated with various concentrations of Met, Cys or l-cysteine (CysH) in the absence or presence of tumor necrosis factor-α (TNF-α). LPL activity was suppressed by TNF-α. In the
absence of TNF-α, Met, Cys and CysH did not change the LPL activity. In the presence of TNF-α, Met and Cys significantly increased
the LPL activity, and Met also enhanced the LPL mRNA level. HSL activity was also suppressed by TNF-α. In the absence of TNF-α,
Met enhanced the HSL activity. In the presence of TNF-α, Met, Cys and CysH suppressed the HSL activity. Sulfur amino acids
such as Met, Cys and CysH affected the LPL activity, mRNA level, and HSL activity in 3T3-L1 adipocytes. Some of these effects
of sulfur amino acids were different between LPL and HSL, between the absence and the presence of TNF-α, and between 3T3-L1
adipocytes and the adipose tissue from rats. 相似文献
12.
脂蛋白脂酶基因的研究进展 总被引:15,自引:3,他引:12
脂蛋白脂酶(lipoprotein lipase, LPL)是脂质代谢的关键酶, 主要催化乳糜微粒和极低密度脂蛋白中的甘油三酯水解, 产生供组织利用的脂肪酸和单酰甘油。LPL基因突变影响LPL活性, 导致脂质代谢紊乱, 与2型糖尿病、高血压、动脉硬化、肥胖、冠心病的发病风险相关联。文章综述了LPL基因的结构、功能、表达调控以及与复杂疾病的关联研究进展。 相似文献
13.
Kirsten Turlo Calvin S. Leung Jane J. Seo Chris N. Goulbourne Oludotun Adeyo Peter Gin Constance Voss André Bensadoun Loren G. Fong Stephen G. Young Anne P. Beigneux 《Biochimica et Biophysica Acta (BBA)/Molecular and Cell Biology of Lipids》2014,1841(7):963-969
The S447X polymorphism in lipoprotein lipase (LPL), which shortens LPL by two amino acids, is associated with low plasma triglyceride levels and reduced risk for coronary heart disease. S447X carriers have higher LPL levels in the pre- and post-heparin plasma, raising the possibility that the S447X polymorphism leads to higher LPL levels within capillaries. One potential explanation for increased amounts of LPL in capillaries would be more avid binding of S447X-LPL to GPIHBP1 (the protein that binds LPL dimers and shuttles them to the capillary lumen). This explanation seems plausible because sequences within the carboxyl terminus of LPL are known to mediate LPL binding to GPIHBP1. To assess the impact of the S447X polymorphism on LPL binding to GPIHBP1, we compared the ability of internally tagged versions of wild-type LPL (WT-LPL) and S447X-LPL to bind to GPIHBP1 in both cell-based and cell-free binding assays. In the cell-based assay, we compared the binding of WT-LPL and S447X-LPL to GPIHBP1 on the surface of cultured cells. This assay revealed no differences in the binding of WT-LPL and S447X-LPL to GPIHBP1. In the cell-free assay, we compared the binding of internally tagged WT-LPL and S447X-LPL to soluble GPIHBP1 immobilized on agarose beads. Again, no differences in the binding of WT-LPL and S447X-LPL to GPIHBP1 were observed. We conclude that increased binding of S447X-LPL to GPIHBP1 is unlikely to be the explanation for more efficient lipolysis and lower plasma triglyceride levels in S447X carriers. 相似文献
14.
《Biochimica et Biophysica Acta (BBA)/Lipids and Lipid Metabolism》1997,1344(2):132-138
Lipoprotein lipase (LPL), an enzyme playing the central role in triglyceride metabolism, is a glycoprotein and a homodimer of identical subunits. Dimerization and proper processing of oligosaccharide chains are important maturation steps in post-translational regulation of enzyme activity. Indirect evidences suggest that dimerization of LPL occurs in endoplasmic reticulum (ER) or Golgi. In this study, we investigated the dimerization status of LPL in 3T3-L1 adipocytes, using sucrose density gradient ultracentrifugation and carbonyl cyanide m-chlorophenylhydrazone (CCCP), an inhibitor of ER-Golgi protein transport. In the presence of CCCP, no increase of cellular LPL activity was detected during 2 h of recovery period after the depletion of LPL with heparin and cycloheximide. Only endoglycosidase H (endo H)-sensitive subunits were found in CCCP-treated cells after endo H digestion, suggesting that inactive LPL was retained in ER. In the presence of castanospermine, an inhibitor of ER glucosidase I, LPL subunits of both control and CCCP-treated cells had same molecular weight, indicating that complete oligosaccharides were transferred to LPL subunits in the presence of CCCP. In sucrose density gradient ultracentrifugation, all the LPL protein synthesized in the presence of CCCP was found at the dimeric fractions as in control cells. Most of LPL protein in control cells showed high affinity for heparin, and there was no difference between the control and CCCP-treated cells. These results suggest that dimerization and acquisition of high affinity for heparin of LPL can occur in ER of CCCP-treated cells without acquisition of catalytic activity. 相似文献
15.
André Bensadoun Charlene D. Mottler Chris Pelletier Daniel Wu Jane J. Seo Calvin S. Leung Oludotun Adeyo Chris N. Goulbourne Peter Gin Loren G. Fong Stephen G. Young Anne P. Beigneux 《Biochimica et Biophysica Acta (BBA)/Molecular and Cell Biology of Lipids》2014,1841(7):970-976
Lipoprotein lipase (LPL) has been highly conserved through vertebrate evolution, making it challenging to generate useful antibodies. Some polyclonal antibodies against LPL have turned out to be nonspecific, and the available monoclonal antibodies (Mabs) against LPL, all of which bind to LPL's carboxyl terminus, have drawbacks for some purposes. We report a new LPL-specific monoclonal antibody, Mab 4-1a, which binds to the amino terminus of LPL (residues 5–25). Mab 4-1a binds human and bovine LPL avidly; it does not inhibit LPL catalytic activity nor does it interfere with the binding of LPL to heparin. Mab 4-1a does not bind to human hepatic lipase. Mab 4-1a binds to GPIHBP1-bound LPL and does not interfere with the ability of the LPL–GPIHBP1 complex to bind triglyceride-rich lipoproteins. Mab 4-1a will be a useful reagent for both biochemists and clinical laboratories. 相似文献
16.
Y H Ma T Bruin S Tuzgol B I Wilson G Roederer M S Liu J Davignon J J Kastelein J D Brunzell M R Hayden 《The Journal of biological chemistry》1992,267(3):1918-1923
We are studying naturally occurring mutations in the gene for lipoprotein lipase (LPL) to advance our knowledge about the structure/function relationships for this enzyme. We and others have previously described 11 mutations in human LPL gene and until now none of these directly involves any of the residues in the proposed Asp156-His241-Ser132 catalytic triad. Here we report two separate probands who are deficient in LPL activity and have three different LPL gene haplotypes, suggesting three distinct mutations. Using polymerase chain reaction cloning and DNA sequencing we have identified that proband 1 is a compound heterozygote for a G----A transition at nucleotide 721, resulting in a substitution of asparagine for aspartic acid at residue 156, and a T----A transversion, resulting in a substitution of serine for cysteine at residues 216. Proband 2 is homozygous for an A----G base change at nucleotide 722, leading to a substitution of glycine for aspartic acid at residue 156. The presence of these mutations in the patients and available family members was confirmed by restriction analysis of polymerase chain reaction-amplified DNA. In vitro site-directed mutagenesis and subsequent expression in COS cells have confirmed that all three mutations result in catalytically defective LPL. The two naturally occurring mutations, which both alter the same aspartic acid residue in the proposed Asp156-His241-Ser132 catalytic triad of human LPL, indicate that Asp156 plays a significant role in LPL catalysis. The Cys216----Ser mutation destroys a conserved disulfide bridge that is apparently critical for maintaining LPL structure and function. 相似文献
17.
Werner J. Geldenhuys Joel Caporoso Thomas C. Leeper Yoon-Kwang Lee Li Lin Altaf S. Darvesh Prabodh Sadana 《Bioorganic & medicinal chemistry letters》2017,27(2):303-308
Elevated triglycerides (TG) contribute towards increased risk for cardiovascular disease. Lipoprotein lipase (LPL) is an enzyme that is responsible for the metabolism of core triglycerides of very-low density lipoproteins (VLDL) and chylomicrons in the vasculature. In this study, we explored the structure-activity relationships of our lead compound (C10d) that we have previously identified as an LPL agonist. We found that the cyclopropyl moiety of C10d is not absolutely necessary for LPL activity. Several substitutions were found to result in loss of LPL activity. The compound C10d was also tested in vivo for its lipid lowering activity. Mice were fed a high-fat diet (HFD) for four months, and treated for one week at 10 mg/kg. At this dose, C10d exhibited in vivo biological activity as indicated by lower TG and cholesterol levels as well as reduced body fat content as determined by ECHO-MRI. Furthermore, C10d also reduced the HFD induced fat accumulation in the liver. Our study has provided insights into the structural and functional characteristics of this novel LPL activator. 相似文献
18.
Lipoprotein lipase (LPL) is a key enzyme in lipid metabolism. Decrease of the LPL enzymatic activity leads to elevated triglycerides (TG) and reduced high-density lipoprotein (HDL-C levels), both risk factors for cardiovascular disease (CVD). Therefore, mutations, which decrease the LPL activity, may confer susceptibility to CVD. Here, the informational spectrum method (ISM), a virtual spectroscopy method for structure/function analysis of nucleotide and protein sequences, is applied for identification of evolutionary highly conserved information encoded by the primary structure of LPL. It was demonstrated that mutations, which alter the LPL enzymatic activity also alter this information. On the basis of this finding, an efficient and simple bioinformatics criterion for assessment of the pathogenic effect of LPL nonsynonymous single nucleotide substitution as a risk factor of CVD has been proposed. 相似文献
19.
Overexpression of the adipocyte differentiation and determination factor-1 (ADD-1) or sterol regulatory element binding protein-1 (SREBP-1) induces the expression of numerous genes involved in lipid metabolism, including lipoprotein lipase (LPL). Therefore, we investigated whether LPL gene expression is controlled by changes in cellular cholesterol concentration and determined the molecular pathways involved. Cholesterol depletion of culture medium resulted in a significant induction of LPL mRNA in the 3T3-L1 preadipocyte cell line, whereas addition of cholesterol reduced LPL mRNA expression to basal levels. Similar to the expression of the endogenous LPL gene, the activity of the human LPL gene promoter was enhanced by cholesterol depletion in transient transfection assays, whereas addition of cholesterol caused a reversal of its induction. The effect of cholesterol depletion upon the human LPL gene promoter was mimicked by cotransfection of expression constructs encoding the nuclear form of SREBP-1a, -1c (also called ADD-1) and SREBP-2. Bioinformatic analysis demonstrated the presence of 3 potential sterol regulatory elements (SRE) and 3 ADD-1 binding sequences (ABS), also known as E-box motifs. Using a combination of in vitro protein-DNA binding assays and transient transfection assays of reporter constructs containing mutations in each individual site, a sequence element, termed LPL-SRE2 (SRE2), was shown to be the principal site conferring sterol responsiveness upon the LPL promoter. These data furthermore underscore the importance of SRE sites relative to E-boxes in the regulation of LPL gene expression by sterols and demonstrate that sterols contribute to the control of triglyceride metabolism via binding of SREBP to the LPL regulatory sequences. 相似文献
20.
Physiological regulation of lipoprotein lipase 总被引:1,自引:0,他引:1
Sander Kersten 《Biochimica et Biophysica Acta (BBA)/Molecular and Cell Biology of Lipids》2014,1841(7):919-933