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1.
The molecular mechanism for the genetic disorder familial defective apolipoprotein B100 总被引:10,自引:0,他引:10
Borén J Ekström U Agren B Nilsson-Ehle P Innerarity TL 《The Journal of biological chemistry》2001,276(12):9214-9218
Familial defective apolipoprotein B100 (FDB) is a genetic disorder in which low density lipoproteins (LDL) bind defectively to the LDL receptor, resulting in hypercholesterolemia and premature atherosclerosis. FDB is caused by a mutation (R3500Q) that changes the conformation of apolipoprotein (apo) B100 near the receptor-binding site. We previously showed that arginine, not simply a positive charge, at residue 3500 is essential for normal receptor binding and that the carboxyl terminus of apoB100 is necessary for mutations affecting arginine 3500 to disrupt LDL receptor binding. Thus, normal receptor binding involves an interaction between arginine 3500 and tryptophan 4369 in the carboxyl tail of apoB100. W4369Y LDL and R3500Q LDL isolated from transgenic mice had identically defective LDL binding and a higher affinity for the monoclonal antibody MB47, which has an epitope flanking residue 3500. We conclude that arginine 3500 interacts with tryptophan 4369 and facilitates the conformation of apoB100 required for normal receptor binding of LDL. From our findings, we developed a model that explains how the carboxyl terminus of apoB100 interacts with the backbone of apoB100 that enwraps the LDL particle. Our model also explains how all known ligand-defective mutations in apoB100, including a newly discovered R3480W mutation in apoB100, cause defective receptor binding. 相似文献
2.
Flood C Gustafsson M Richardson PE Harvey SC Segrest JP Borén J 《The Journal of biological chemistry》2002,277(35):32228-32233
An initial event in atherosclerosis is the retention of lipoproteins within the intima of the vessel wall. Previously we identified Site B (residues 3359-3369) in apolipoprotein (apo) B100 as the proteoglycan binding sequence in low density lipoproteins (LDLs) and showed that the atherogenicity of apoB-containing lipoproteins is linked to their affinity for artery wall proteoglycans. However, both apoB100- and apoB48-containing lipoproteins are equally atherogenic even though Site B lies in the carboxyl-terminal half of apoB100 and is absent in apoB48. If binding to proteoglycans is a key step in atherogenesis, apoB48-containing lipoproteins must bind to proteoglycans via other proteoglycan binding sites in the amino-terminal 48% of apoB. In vitro studies have identified five clusters of basic amino acids in delipidated apoB48 that bind negatively charged glycosaminoglycans. To determine which of these sites is functional on LDL particles, we analyzed the proteoglycan binding activity of recombinant human LDLs from transgenic mice or rat hepatoma cells. Substitution of neutral amino acids for the basic amino acids in Site B-Ib (residues 84-94) abolished the proteoglycan binding activity of recombinant apoB53. Carboxyl-truncated apoB80 bound biglycan with higher affinity than apoB100 and apoB48. ApoB80 in which Site B was mutated had the same affinity for proteoglycans as apoB48. These data support the hypothesis that the carboxyl terminus of apoB100 "masks" Site B-Ib, the amino-terminal proteoglycan binding site, and that this site is exposed in carboxyl-truncated forms of apoB. The presence of a proteoglycan binding site in the amino-terminal region of apoB may explain why apoB48- and apoB100-containing lipoproteins are equally atherogenic. 相似文献
3.
Conformation of apolipoprotein B after lipid extraction of low density lipoproteins attached to an electron microscope grid 总被引:4,自引:0,他引:4
We have examined the shape of apolipoprotein B (apoB) from low density lipoproteins (LDL) using a new method to prepare the electron microscope grids. After adsorption of the lipoproteins to a carbon-coated copper grid, lipids were extracted with ethanol-ether 4:1; an aqueous negative stain was then applied. When the LDL residue was examined after this treatment, apoB, together with residual lipid, appeared as an elongated flexible structure about 600-700 A in length consisting of multiple domains of variable width from 20-70 A. Occasionally, the elongated apoB formed an irregular ring-shaped structure, but most of the rings were open. When LDL were pretreated with glutaraldehyde, then adsorbed, extracted, and stained, most of the images were closed rings with an average contour length of 700 A, again consisting of multiple domains of variable sizes. These results are consistent with apoB being composed of multiple domains arranged in an elongated structure on the surface of the LDL, and with distant domains possessing a mutual affinity that favors their cross-linking. 相似文献
4.
Locating a low density lipoprotein-targeting domain of human apolipoprotein B-100 by expressing a minigene construct in transgenic mice 总被引:2,自引:0,他引:2
W J Xiong E Zsigmond A M Gotto K Y Lei L Chan 《The Journal of biological chemistry》1991,266(31):20893-20898
Through its interaction with the low density lipoprotein (LDL) receptor, apolipoprotein (apo) B-100 is a major determinant of LDL metabolism and plasma cholesterol. Its receptor binding ability is conformation-dependent and requires its expression on the right lipoprotein particles. The structural signal that targets apoB-100 to LDL is unknown. We have microinjected a human apoB-100 minigene construct comprising less than 25% of the apoB-100 sequence driven by the natural apoB promoter to produce transgenic mice. The transgene product was expressed at a high level and was present exclusively in the LDL of these animals. Analysis of the responsible sequence (residues 2878-3925 of apoB-100) reveals unique structural features that may be important in its role as an LDL-targeting domain. 相似文献
5.
Garner B Harvey DJ Royle L Frischmann M Nigon F Chapman MJ Rudd PM 《Glycobiology》2001,11(10):791-802
The carbohydrate composition of apolipoprotein (apo) B100, particularly its degree of sialylation, may contribute to the atherogenic properties of low-density lipoprotein (LDL). We analyzed LDL apoB100 glycans derived from normolipidemic, hypercholesterolemic, and hypertriglyceridemic diabetic subjects. Using exoglycosidase carbohydrate sequencing and matrix-assisted laser desorption/ionization mass spectrometry to analyze fluorescently labeled oligosaccharides, we report evidence for several carbohydrates not previously identified on apoB100, including truncated complex biantennary N-glycans and hybrid N-glycans. The distribution and diversity of the apoB100 glycans isolated from all individuals was highly conserved. The N-glycan composition of apoB100 derived from five LDL subpopulations (LDL1, d = 1.018-1.023; LDL2, d = 1.023-1.030; LDL3, d = 1.030-1.040; LDL4, d = 1.040-1.051; LDL5, d = 1.051-1.065 g/ml) did not vary in normolipidemic or hypercholesterolemic subjects. Furthermore, we found no evidence for "desialylated" apoB100 glycans in any of the samples analyzed. Analysis of the most abundant LDL ganglioside, alpha-N-acetylneuraminyllactosyl-ceramide, revealed a deficiency in small dense LDL and in the most buoyant subpopulation. These data provide a novel explanation for the apparent deficiency of sialic acid in small dense LDL and indicate that the global apoB100 N-glycan composition is invariable in the patient groups studied. 相似文献
6.
Arterial retention of apolipoprotein B(48)- and B(100)-containing lipoproteins in atherogenesis 总被引:5,自引:0,他引:5
PURPOSE OF REVIEW: The "response to retention" hypothesis of atherosclerosis suggests that the arterial deposition of cholesterol is directly proportional to the concentration of circulating plasma lipoproteins. However, there is increasing evidence to support the concept that specific lipoproteins may be preferentially retained within the arterial wall, possibly as a result of greater affinity for cell surface and extracellular matrices. RECENT FINDINGS: Recently, key studies have provided insight into mechanisms involved in the interaction of apolipoprotein B (apoB)-containing lipoproteins with extracellular matrices. In addition, novel methods and innovative experimental design has enabled us to differentiate between the delivery, retention and efflux of apoB(48)- and apoB(100)-containing lipoproteins. Other studies have demonstrated a relationship between extracellular matrix proteoglycan expression and the development of atherosclerosis. Discussion in the present review also extends to the mechanisms that are involved in the relative intimal retention of apoB(48)- and apoB(100)-containing lipoproteins in order to explain the atherogenicity of these macromolecules. SUMMARY: The perspective of this review is to highlight recent advances in the area of arterial lipoprotein retention and the physiological significance these processes may have in the aetiology of cardiovascular disease. Importantly, an understanding of the mechanisms responsible for the retention of apoB(48)/B(100)-containing lipoproteins will enable new strategies to be developed for the future management of cardiovascular disease. 相似文献
7.
De novo production of low density lipoproteins: fact or fancy 总被引:2,自引:0,他引:2
Many investigators, observing an apparent dilution in the plasma specific activity (SA) of apolipoprotein B-100 (apoB) in low density lipoprotein (LDL) as compared with that in very low density lipoprotein (VLDL) after injection of radiolabeled VLDL, have formulated kinetic hypotheses incorporating the concept of de novo production of LDL to explain their data in humans and other mammals. These hypotheses, with rare exception, do not account for the kinetic heterogeneity known to exist in the apoB of human VLDL on the basis of size and in the apoB of rabbit VLDL on the basis of size and presence of apolipoprotein E. When a logical analysis of such kinetic heterogeneity of apoB in plasma VLDL is performed, it becomes clear that the apparent dilution of the SA of apoB in LDL relative to that in VLDL can be explained without the requirement for de novo production of LDL. Although this alternative hypothesis, incorporating the concept of kinetic heterogeneity of apoB in VLDL, does not exclude the process of de novo production of LDL, which so many investigators have invoked to explain their data, it does raise a question as to the existence of such a process since an alternative hypothesis can explain such data just as well. Clearly, more experimental data on the kinetic heterogeneity of human and other mammalian VLDL are needed before a reasonable choice can be made between these two hypotheses. 相似文献
8.
A Leroy G Castro G Agnani R Sa?le A Barkia J C Fruchart 《Journal of lipid research》1992,33(6):889-898
Rabbit plasma low density lipoprotein (LDL) contains one major apolipoprotein of apparent molecular weight of 320 kDa, designated apolipoprotein (apo) Bh, while another component termed apoB1 of apparent molecular weight of 220 kDa is found in chylomicrons. The fragments generated by thrombin digestion of the protein moieties of rabbit and human LDL were separated by polyacrylamide gradient gel electrophoresis and compared. As in the human species, the enzyme produced limited cleavage patterns of rabbit LDL apoB. Within the first 2 h, two fragments (Tr1 and Tr2, with apparent molecular weights 280,000 and 44,000, respectively) appeared. Longer incubations led to the production of two additional peptides, Tr3 and Tr4 (apparent molecular weights 180,000 and 96,000, respectively). Ten monoclonal antibodies, developed against rabbit LDL and designated P01 to P10, were found to react with rabbit apoB. Some also cross-reacted with human apoB. Epitope mapping, performed with these antibodies, showed that Tr3 and Tr4 were derived from the further degradation of Tr1. The rabbit is one of the most frequently used animals in atherosclerosis research. Its LDL receptor has been characterized and there exists a strain of homozygous LDL receptor-deficient rabbits referred to as WHHL rabbits. Despite this, little has been done to characterize the structure of rabbit apoB; only a short region has been sequenced and shown to be the carboxyl-terminal region, the rabbit apoB1. The molecular weight of human apoB (550,000) is much larger than rabbit apoBh. In both species, a primary and secondary thrombin cleavage occur, but the size of the fragments produced is very different between the two species. Identification of the thrombolytic fragments of the rabbit apoB have afforded the opportunity to compare the structures of both apoB species. 相似文献
9.
Molecular basis of the interaction between plasma platelet-activating factor acetylhydrolase and low density lipoprotein 总被引:5,自引:0,他引:5
Stafforini DM Tjoelker LW McCormick SP Vaitkus D McIntyre TM Gray PW Young SG Prescott SM 《The Journal of biological chemistry》1999,274(11):7018-7024
The platelet-activating factor acetylhydrolases are enzymes that were initially characterized by their ability to hydrolyze platelet-activating factor (PAF). In human plasma, PAF acetylhydrolase (EC 3.1.1.47) circulates in a complex with low density lipoproteins (LDL) and high density lipoproteins (HDL). This association defines the physical state of PAF acetylhydrolase, confers a long half-life, and is a major determinant of its catalytic efficiency in vivo. The lipoprotein-associated enzyme accounts for all of the PAF hydrolysis in plasma but only two-thirds of the protein mass. To characterize the enzyme-lipoprotein interaction, we employed site-directed mutagenesis techniques. Two domains within the primary sequence of human PAF acetylhydrolase, tyrosine 205 and residues 115 and 116, were important for its binding to LDL. Mutation or deletion of those sequences prevented the association of the enzyme with lipoproteins. When residues 115 and 116 from human PAF acetylhydrolase were introduced into mouse PAF acetylhydrolase (which normally does not associate with LDL), the mutant mouse PAF acetylhydrolase associated with lipoproteins. To analyze the role of apolipoprotein (apo) B100 in the formation of the PAF acetylhydrolase-LDL complex, we tested the ability of PAF acetylhydrolase to bind to lipoproteins containing truncated forms of apoB. These studies indicated that the carboxyl terminus of apoB plays a key role in the association of PAF acetylhydrolase with LDL. These data on the molecular basis of the PAF acetylhydrolase-LDL association provide a new level of understanding regarding the pathway for the catabolism of PAF in human blood. 相似文献
10.
Radical reaction of low-density lipoprotein (LDL) is a key step in atherogenesis and causes both a decrease in the sialic acid moiety and modification of apolipoprotein B-100 (apoB). Although apoB modification (cross-link and fragmentation) increases in atherosclerosis, the change in apoB-bound sialic acid in atherosclerosis is controversial. To elucidate the physiological implications of desialylation of LDL by radical reaction, the reactivity of sialic acid of LDL was compared with that of apoB, which underwent facile fragmentation in radical reactions. ApoB was determined by immunoblot analysis with anti-apoB antiserum, and the sialic acid moiety was measured by blot analysis with a biotin-bound lectin [biotin-SSA from Japanese elderberry (Sambucus sieboldiana)] specific to sialic acid. When human LDL was oxidized with Cu(2+) at 37 degrees C, apoB and apoB-attached sialic acid decreased simultaneously. Comparison of the staining bands with anti-apoB and with biotin-SSA shows that sialic acid moieties still remain on fragmented apoB proteins, indicating that the decrease in sialic acid is much slower than that of apoB fragmentation. In addition, human plasma was oxidized with 400 microM of Cu(2+) at 37 degrees C. Similar analysis indicates that the decrease in sialic acid attached to apoB also results from the fragmentation of apoB. This study indicates that the fragmentation of apoB proceeds at a much faster rate than the decrease in sialic acid content when a free radical reaction is induced in isolated LDL as well as in plasma LDL exposed to Cu(2+)-induced oxidative stress. On the basis of these results, the modification of apoB is much more sensitive than the decrease in sialic acid as an indicator of oxidative stress. 相似文献
11.
The apoprotein (apoB) of low density lipoprotein (LDL) is reported to be a large polypeptide, and it is proposed that there are two similar-sized subunit proteins in LDL (Smith, Dawson, and Tanford. 1972. J. Biol. Chem. 247: 3376-3381.). When apoB is isolated under conditions that minimize artifactual proteolysis, only a single, large molecular weight protein appears on polyacrylamide gel electrophoresis in SDS. To investigate the organization of apoB as it exists within native LDL, limited proteolysis with trypsin has been used as a structural probe. Tryptic digestion for 1 hr at pH 7.6 with enzyme-to-protein ratios of 1:100 and 1:5 results in the liberation of approximately 10% and 30% of apoB as smaller, water-soluble peptides. These peptides may be separated from the partially digested but still intact tryptic core (T-core) of the lipoprotein by chromatography on Sephadex G-75. Repeatedly, the 1:5 T-core of native LDL is found to contain a family of polypeptides of 14,000-100,000 molecular weight. Although they have lost significant quantities of apoprotein, these T-cores sustain an appearance of homogeneity, as studied by analytical ultracentrifugation. Their measured molecular weights do not differ appreciably from those of the native LDL, and the carbohydrate content of the 1:5 tryptic T-core of LDL is similar to that of the native LDL. In normolipemic individuals, LDL generally exists in a monodisperse state, but, in different individuals, monodisperse LDL may range in molecular weight from 2.4 to 3.9 x 10(6). Limited tryptic digestions were used to probe the organization of apoB in these different molecular weight LDL. As assayed by SDS-acrylamide gel electrophoresis of the larger polypeptides and fingerprinting of the smaller released peptides, those regions of LDL exposed to trypsin digestion are identical in monodisperse LDL of 2.5 and 3.4 x 10(6) molecular weight. Thus, the different quantities of lipid bound in these various LDL must interact with apoB so that the same regions of the apoprotein are exposed to the action of trypsin in these different molecular weight lipoproteins. 相似文献
12.
Effect of cell cholesterol content on apolipoprotein B secretion and LDL receptor activity in the human hepatoma cell line, HepG2 总被引:1,自引:0,他引:1
Human hepatoma HepG2 cells were used to study the effects of cholesterol loading and depletion on apolipoprotein B (apoB) secretion and low-density lipoprotein (LDL) receptor activity. Exposure of HepG2 cells to cholesterol and oleic acid, which elevated intracellular cholesterol levels, stimulated apoB secretion and reduced receptor-mediated uptake of LDL, whereas recombinant complexes of apolipoprotein A-I with dimyristoylphosphatidylcholine, which depleted the cellular cholesterol pool, inhibited apoB secretion and up-regulated LDL receptors. Significant negative correlation (r = -0.92, P less than 0.001) between the levels of apoB secretion and LDL uptake was found. These data suggest that the cholesterol content of the cells may induce concomitant changes in apoB secretion and LDL receptor activity. 相似文献
13.
Krisko A Stjepanović G Pifat G Ruysschaert JM Goormaghtigh E 《Biochimica et biophysica acta》2007,1768(11):2923-2930
Conformational changes of human plasma apolipoprotein B100 (apoB) during oxidative modification of low-density lipoproteins (LDL) have been investigated. Emphasis has been put on the early stages of LDL oxidation and the modification of apoB. We have applied two different modes of LDL oxidation initiation in order to approach the problem from different perspectives. To study conformational changes of the protein and the phospholipids surface monolayer, we have applied attenuated total reflection infrared as well as fluorescence spectroscopy. We have found for the first time that conformational changes of apoB occur even in the earliest stages of oxidation process and that those are located predominantly in the beta-sheet regions. The dynamics of changes has also been described and related to different stages of oxidation. After initial increase in particle surface accessibility and mobility, by entering into the propagation phase of oxidation process, LDL surface accessibility and mobility are decreased. Finally, in the decomposition phase of LDL oxidation, as the particle faces large chemical and physical changes, surface mobility and accessibility is increased again. These observations provide new insights into the modifications of LDL particles upon oxidation. 相似文献
14.
Conformational changes of human plasma apolipoprotein B100 (apoB) during oxidative modification of low-density lipoproteins (LDL) have been investigated. Emphasis has been put on the early stages of LDL oxidation and the modification of apoB. We have applied two different modes of LDL oxidation initiation in order to approach the problem from different perspectives. To study conformational changes of the protein and the phospholipids surface monolayer, we have applied attenuated total reflection infrared as well as fluorescence spectroscopy. We have found for the first time that conformational changes of apoB occur even in the earliest stages of oxidation process and that those are located predominantly in the β-sheet regions. The dynamics of changes has also been described and related to different stages of oxidation. After initial increase in particle surface accessibility and mobility, by entering into the propagation phase of oxidation process, LDL surface accessibility and mobility are decreased. Finally, in the decomposition phase of LDL oxidation, as the particle faces large chemical and physical changes, surface mobility and accessibility is increased again. These observations provide new insights into the modifications of LDL particles upon oxidation. 相似文献
15.
Structure of low density lipoprotein (LDL) particles: basis for understanding molecular changes in modified LDL 总被引:12,自引:0,他引:12
Hevonoja T Pentikäinen MO Hyvönen MT Kovanen PT Ala-Korpela M 《Biochimica et biophysica acta》2000,1488(3):189-210
Low density lipoprotein (LDL) particles are the major cholesterol carriers in circulation and their physiological function is to carry cholesterol to the cells. In the process of atherogenesis these particles are modified and they accumulate in the arterial wall. Although the composition and overall structure of the LDL particles is well known, the fundamental molecular interactions and their impact on the structure of LDL particles are not well understood. Here, the existing pieces of structural information on LDL particles are combined with computer models of the individual molecular components to give a detailed structural model and visualization of the particles. Strong evidence is presented in favor of interactions between LDL lipid constituents that lead to specific domain formation in the particles. A new three-layer model, which divides the LDL particle into outer surface, interfacial layer, and core, and which is capable of explaining some seemingly contradictory interpretations of molecular interactions in LDL particles, is also presented. A new molecular interaction model for the beta-sheet structure and phosphatidylcholine headgroups is introduced and an overall view of the tertiary structure of apolipoprotein B-100 in the LDL particles is presented. This structural information is also utilized to understand and explain the molecular characteristics and interactions of modified, atherogenic LDL particles. 相似文献
16.
Direct determination of human and rabbit apolipoprotein B selectively precipitated with butanol-isopropyl ether 总被引:1,自引:0,他引:1
A method is described for the rapid, selective, and quantitative precipitation of apolipoprotein B from isolated hypercholesterolemic rabbit and human very low density lipoproteins (VLDL), intermediate density lipoproteins (IDL), and low density lipoproteins (LDL). Lipoprotein samples are heat-treated at 100 degrees C in 1% SDS. The denatured apoprotein solutions are then mixed briefly with two volumes of butanol-isopropyl ether 45:55 (v/v) to precipitate the apoB. The supernatant solutions, containing the non-apoB proteins and lipids, are removed and the apoB pellet is washed once with water. To determine apoB specific activity, the apoB pellet is resolubilized in 0.5 M NaOH by heating for 30 min at 120 degrees C. The hydrolyzed apoB protein is quantitated by fluorescence of a fluorescamine derivative. The precipitation of apoB is quantitative and selective: 99.5% of rabbit 125I-labeled LDL-apoB and 97.5% of human 125I-labeled LDL-apoB is precipitated and less than 5% of 125I-labeled HDL added to unlabeled VLDL, IDL, or LDL is precipitated. Triglyceride and cholesteryl ester contamination of the apoB pellet is less than 2% of their original radioactivities. 相似文献
17.
Bourdon E Loreau N Lagrost L Davignon J Bernier L Blache D 《Free radical biology & medicine》2006,41(5):786-796
Oxidized low-density lipoproteins (Ox-LDL) are key elements in atherogenesis. Apolipoprotein AI (apoAI) is an active component of the antiatherogenic high-density lipoproteins (HDL). In contrast, plasma apolipoprotein B (apoB), the main component of LDL, is highly correlated with coronary risk. Our results, obtained in HepG2 cells, show that Ox-LDL, unlike native LDL, leads to opposite effects on apoB and apoAI, namely a decrease in apoAI and an increase in apoB secretion as evaluated by [(3)H]leucine incorporation and specific immunoprecipitation. Parallel pulse-chase studies show that Ox-LDL impaired apoB degradation, whereas apoAI degradation was increased and mRNA levels were decreased. We also found that enhanced lipid biosynthesis of both triglycerides and cholesterol esters was involved in the Ox-LDL-induced increase in apoB secretion. Our data suggest that the increase in apoB and decrease in apoAI secretion may in part contribute to the known atherogenicity of Ox-LDL through an elevated LDL/HDL ratio, a strong predictor of coronary risk in patients. 相似文献
18.
Interactions of low density lipoprotein2 and other apolipoprotein B-containing lipoproteins with lipoprotein(a) 总被引:2,自引:0,他引:2
S Q Ye V N Trieu D L Stiers W J McConathy 《The Journal of biological chemistry》1988,263(13):6337-6343
Studies were undertaken to investigate potential interactions among plasma lipoproteins. Techniques used were low density lipoprotein2 (LDL2)-ligand blotting of plasma lipoproteins separated by nondenaturing 2.5-15% gradient gel electrophoresis, ligand binding of plasma lipoproteins by affinity chromatography with either LDL2 or lipoprotein(a) (Lp(a)) as ligands, and agarose lipoprotein electrophoresis. Ligand blotting showed that LDL2 can bind to Lp(a). When apolipoprotein(a) was removed from Lp(a) by reduction and ultracentrifugation, no interaction between LDL2 and reduced Lp(a) was detected by ligand blotting. Ligand binding showed that LDL2-Sepharose 4B columns bound plasma lipoproteins containing apolipoproteins(a), B, and other apolipoproteins. The Lp(a)-Sepharose column bound lipoproteins containing apolipoprotein B and other apolipoproteins. Furthermore, the Lp(a) ligand column bound more lipoprotein lipid than the LDL2 ligand column, with the Lp(a) ligand column having a greater affinity for triglyceride-rich lipoproteins. Lipoprotein electrophoresis of a mixture of LDL2 and Lp(a) demonstrated a single band with a mobility intermediate between that of LDL2 and Lp(a). Chemical modification of the lysine residues of apolipoprotein B (apoB) by either acetylation or acetoacetylation prevented or diminished the interaction of LDL2 with Lp(a), as shown by both agarose electrophoresis and ligand blotting using modified LDL2. Moreover, removal of the acetoacetyl group from the lysine residues of apoB by hydroxylamine reestablished the interaction of LDL2 with Lp(a). On the other hand, blocking of--SH groups of apoB by iodoacetamide failed to show any effect on the interaction between LDL2 and Lp(a). Based on these observations, it was concluded that Lp(a) interacts with LDL2 and other apoB-containing lipoproteins which are enriched in triglyceride; this interaction is due to the presence of apolipoprotein(a) and involves lysine residues of apoB interacting with the plasminogen-like domains (kringle 4) of apolipoprotein(a). Such results suggest that Lp(a) may be involved in triglyceride-rich lipoprotein metabolism, could form transient associations with apoB-containing lipoproteins in the vascular compartment, and alter the intake by the high affinity apoB, E receptor pathway. 相似文献
19.
M Sorci-Thomas M D Wilson F L Johnson D L Williams L L Rudel 《The Journal of biological chemistry》1989,264(15):9039-9045
African green monkeys were fed diets containing low and moderate cholesterol concentrations with either polyunsaturated or unsaturated fat as 40% of calories. Plasma total cholesterol, low density lipoprotein (LDL) cholesterol, and apoB concentrations generally were higher in animals fed (a) the higher dietary cholesterol concentration and (b) saturated fat. At necropsy, liver and intestine were removed, and measurement of mRNAs for LDL receptors (liver) and for apolipoprotein B (liver and intestine) was done. Monkey small intestine mucosa made exclusively apoB48 while the liver made only apoB100, although apoB mRNA in both tissues was the same size (14 kilobases). No dietary cholesterol or fat effects were found for apoB mRNA abundance in the liver, while the animals fed the higher dietary cholesterol level had 50% lower levels of hepatic LDL receptor mRNA. In a separate group of animals, livers were perfused and the rate of apoB secretion was measured. No dietary fat effect on apoB secretion rate was found, and no relationship between plasma LDL cholesterol concentration and the rate of hepatic apoB production existed. These findings support the idea that the dietary factors that increase LDL concentrations act by reducing clearance of apoB-containing particles rather than by increasing production of these lipoproteins. Hepatic LDL receptor mRNA was similar in abundance in polyunsaturated fat and saturated fat-fed animals, suggesting that the difference in plasma cholesterol concentration between these groups is not mediated via effects on LDL receptor mRNA abundance. The level of intestinal apoB mRNA was about 30% higher in animals fed the moderate dietary cholesterol concentration. Earlier studies have shown that more cholesterol is transported in chylomicrons from the intestine when dietary cholesterol levels are higher, and the increased intestinal apoB mRNA abundance may reflect increased intestinal cholesterol transport and chylomicron apoB48 production. 相似文献
20.
The use of monoclonal antibodies to localize the low density lipoprotein receptor-binding domain of apolipoprotein B 总被引:10,自引:0,他引:10
R Milne R Théolis R Maurice R J Pease P K Weech E Rassart J C Fruchart J Scott Y L Marcel 《The Journal of biological chemistry》1989,264(33):19754-19760
Human apolipoprotein (apo) B-100 is composed of 4536 amino acids. It is thought that the binding of apoB to the low density lipoprotein (LDL) receptor involves an interaction between basic amino acids of the ligand and acidic residues of the receptor. Three alternative models have been proposed to describe this interaction: 1) a single region of apoB is involved in receptor binding; 2) groups of basic amino acids from throughout the apoB primary structure act in concert in apoB receptor binding; and 3) apoB contains multiple independent binding regions. We have found that monoclonal antibodies (Mabs) specific for a region that spans a thrombin cleavage site at apoB residue 3249 (T2/T3 junction) totally blocked LDL binding to the LDL receptor. Mabs specific for epitopes outside this region had either no or partial ability to block LDL binding. In order to define the region of apoB directly involved in the interaction with the LDL receptor we have tested 22 different Mabs for their ability to bind to LDL already fixed to the receptor. A Mab specific for an epitope situated between residues 2835 and 2922 could bind to its epitope on LDL fixed to its receptor whereas a second epitope between residues 2980 and 3084 is inaccessible on receptor-bound LDL. A series of epitopes near residue 3500 of apoB is totally inaccessible, and another situated between residues 4027 and 4081 is poorly accessible on receptor-bound LDL. In contrast, an epitope that is situated between residues 4154 and 4189 is fully exposed. Mabs specific for epitopes upstream and downstream of the region 3000-4000 can bind to receptor-bound LDL with a stoichiometry close to unity. Our results strongly suggest that the unique region of apoB directly involved in the LDL-receptor interaction is that of the T2/T3 junction. 相似文献