Biomedical subjects

S O Olofsson

Publications and source records attributed to S O Olofsson.

At least 19 recordsLinked to original sources

PubMed1992

The assembly and secretion of ApoB 100-containing lipoproteins in Hep G2 cells. ApoB 100 is cotranslationally integrated into lipoproteins.

The possibility that apoB 100 is cotranslationally translocated to the endoplasmic reticulum lumen and integrated into lipoproteins has been investigated. ApoB 100 nascent polypeptides were shown to be secreted from pulse-labeled Hep G2 cells after treatment with puromycin and chase for 1 or 2 h in the presence of puromycin and cycloheximide. These nascent polypeptides banded during sucrose gradient ultracentrifugation between the position of the high (HDL) and the low (LDL) density lipoproteins, revealing an inverse relationship between the length of the polypeptide and the density of the fraction. ApoB 100 occurred in the position of LDL and very low density lipoproteins (VLDL). Electronmicroscopy studies of the apoB-containing particles from the gradient indicated an increase in size with increasing length of the polypeptide. Furthermore, labeling studies indicated that the triglyceride load increased with the length of the polypeptide. An inverse relationship between the size of C-terminally truncated apoB polypeptides and the density of the assembled lipoproteins was also observed in experiments with transfected minigenes coding for apoB 41, apoB 29, and apoB 23. These proteins appeared on HDL particles. Pulse-chase experiments indicated that 80-200-kDa apoB nascent polypeptides on particles with HDL density, with time, were converted into larger polypeptides on lighter particles, to be fully replaced by apoB 100 on LDL-VLDL particles. The formation of these LDL-VLDL particles could be blocked by cycloheximide. Sixty-five percent of pulse-labeled apoB nascent polypeptides present in the microsomal fraction was released by sodium carbonate treatment, and 77% of these polypeptides could be recovered on the immature particles (banding between HDL and LDL) after sucrose gradient ultracentrifugation. Pulse-chase experiments indicated that these nascent polypeptides, on the immature lipoproteins, had the capacity to be precursors for all the apoB 100-containing LDL and VLDL particles formed in the cell. The obtained results indicate that a major portion of the apoB nascent polypeptides in the cell form lipoproteins cotranslationally during the translocation to the lumen of the endoplasmic reticulum.

Apolipoprotein B-100↗

PubMed1992

Effects of growth hormone on apolipoprotein-B (apoB) messenger ribonucleic acid editing, and apoB 48 and apoB 100 synthesis and secretion in the rat liver.

Apolipoprotein-B 48 (apoB 48) and apoB 100 expression and the editing of apoB mRNA have previously been shown to be hormonally regulated in rat liver. We have investigated the effects of hypophysectomy and replacement therapy with T4, cortisol (C), and GH in vivo on the proportion of edited apoB mRNA in rat liver and cultured rat hepatocytes as well as the synthesis and secretion of apoB 48 and apoB 100 in cultured rat hepatocytes. Hypophysectomy decreased the proportion of edited apoB mRNA in intact liver from 62% in normal rats to 29% in hypophysectomized rats. Treatment of hypophysectomized rats with T4 and C did not influence the proportion of edited apoB mRNA, whereas treatment with GH, either alone or together with T4 and C, increased the proportion of edited apoB mRNA to the levels observed in normal rats. In cultured hepatocytes isolated from normal rats, the proportion of apoB 48 (percentage of total labeled apoB) was 78% and decreased to 40% in cells isolated from hypophysectomized rats. Treatment of hypophysectomized rats with T4 and C had no effect on the proportion of apoB 48 present in isolated cells, whereas it increased to 60% after treatment with GH together with T4 and C. The proportion of apoB 48 in the medium was affected by hypophysectomy and the various hormonal treatments in a similar way to that observed in the cells. Results from in vivo labeling experiments suggested that GH alone had the capacity to increase the percentage of apoB 48 in hypophysectomized rats. On the contrary, T4 and C was needed, in addition to GH, to increase the proportion of apoB 48 in isolated hepatocytes from hypophysectomized rats. Our results suggest that this discrepancy is due to a difference between the effect of GH alone on apoB mRNA editing in the intact liver and that in isolated hepatocytes. The total secretion of apoB into the cell culture medium was not affected by hypophysectomy and hormonal treatments of the rats. In conclusion, these results indicate that GH is involved in the regulation of editing of apoB mRNA and the proportion of apoB 48 synthesized and secreted in rat liver. Thus, our observations emphasize the importance of GH as a regulator of lipoprotein metabolism.

Animals↗

PubMed1991

Molecular parameters that control the association of low density lipoprotein apo B-100 with chondroitin sulphate.

The association of low density lipoprotein (LDL) with proteoglycans of the arterial intima, in particular chondroitin 6-sulphate proteoglycans, may contribute to LDL accumulation during atherogenesis. We studied the interactions of apolipoprotein B-100 (apo B-100) peptide segments and model peptides with chondroitin 6-sulphate. The ability of these peptides to inhibit complex formation between LDL and chondroitin 6-sulphate was used as a measurement of the interaction. Results from earlier studies suggest that surface located segments of apo B-100 are responsible for the interaction of LDL with heparin and chondroitin sulphate-rich arterial proteoglycans. Therefore 16 hydrophilic apo B-100 peptides were selected for studies and synthesized with a peptide synthesizer. These synthetic peptides were 7 to 26 amino acids long. Four of the peptides inhibited the association of LDL with chondroitin 6-sulphate, namely apo B segments 4230-4254, 3359-3377, 3145-3157 and 2106-2121. The 3359-3377 segment was the most efficient. A common feature between the interacting peptides was an excess of positively charged side chains and based on these results we synthesized nine model peptides that shared sequence characteristics with the interacting apo B-100 peptides. Five of these: RSGRKRSGK, RSSRKRSGK, RGGRKRGGK, RSRSRSRSR and RGRGRGRGR were shown to block the LDL-chondroitin-6-sulphate association, RSRSRSRSR being the most effective. The results suggest that the optimal association of the peptides with chondroitin 6-sulphate is obtained with a minimal chain length of nine amino acids and a minimum of five positive charges and that flexibility in the binding region is important.

Amino Acid Sequence↗

PubMed1991

Growth hormone regulation of serum lipoproteins in the rat: different growth hormone regulatory principles for apolipoprotein (apo) B and the sexually dimorphic apo E concentrations.

Growth hormone (GH) regulation of serum lipoproteins and apolipoproteins was studied using hypophysectomized (Hx) male and female Sprague-Dawley rats. Hypophysectomies were performed at 45 or 50 days of age. Hx rats were given replacement therapy with L-thyroxine (10 micrograms/kg/d) and hydrocortisone (400 micrograms/kg/d) unless otherwise specified. Bovine GH (bGH) was given either as two daily subcutaneous (SC) injections at 12-hour intervals or as a continuous SC infusion. Serum cholesterol and apolipoprotein (apo) E concentrations decreased after Hx of female rats. In contrast, Hx of male rats resulted in increased serum cholesterol concentrations and had no effect on serum apo E concentrations. There were no effects of Hx on high-density lipoprotein (HDL) apo E levels in male rats in contrast to female rats. bGH given twice daily to Hx male rats had no effect on HDL apo E levels, but a continuous infusion of bGH resulted in a marked increase in HDL apo E concentration, to levels above those of intact male rats. As previously observed in female rats, serum and HDL apo A-I concentrations decreased and serum and low-density lipoprotein (LDL) concentrations of apo B increased after Hx of male rats. Treatment with L-thyroxine and hydrocortisone reduced the serum concentrations of apo B. bGH given alone resulted in even lower concentrations of apo B. Serum concentrations of cholesterol and apo E were unaffected by replacement therapy with L-thyroxine and hydrocortisone. Treatment with bGH alone had similar effects on serum cholesterol, apo E, and apo B concentrations as treatment with L-thyroxine, hydrocortisone, and bGH in combination.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1991

The molecular mechanism for the assembly and secretion of ApoB-100-containing lipoproteins.

We have reviewed the literature on the intracellular transport of ApoB-100 and the assembly of the ApoB-100-containing lipoproteins. ApoB-100 is a large molecule (4536 aa) that requires some 15 min to be completed. During the synthesis, the protein could take one of two pathways: a degradational pathway and a pathway that leads to secretion of the protein on mature lipoproteins. The degradational pathway starts with a cotranslational incorporation of ApoB-100 into the membrane of the endoplasmic reticulum in such a way that a relatively large portion of the sequence is exposed on the cytoplasmic surface of this membrane. The membrane bound ApoB-100 is retained in the ER and will eventually undergo intracellular degradation. To enter the pathway that leads to lipoprotein formation, ApoB-100 has to be cotranslationally translocated to the lumen of the ER. ApoB-100 will interact with the lipids during this translation-translocation process and the mature lipoprotein is released into the lumen of the secretory pathway when ApoB-100 is completed and leaves the ribosome. In addition to the mature lipoproteins, the secretory pathway contains an ApoB-100-containing lipoprotein with the density of a HDL particle. This particle is not secreted from the cells but is retained and eventually degraded. Of importance for the retention are sequences present in the C-terminal half of the protein. The mature lipoproteins rapidly leave the ER lumen and are transported to the Golgi apparatus, through which transfer takes considerably longer. The assembly process is a potential site for the regulation of the secretion of the ApoB-100-containing lipoproteins. This process is dependent on active synthesis of phosphatidylcholine and it is also highly dependent on the rate of triacylglycerol synthesis. On the other hand, ApoB-100 appears to be constitutively expressed. An increase in the rate of lipoprotein assembly induced by an increased triacylglycerol synthesis gives rise to an increased recruitment of ApoB-100 nascent polypeptides to interact cotranslationally with lipids. ApoB-100 that is not used for lipoprotein assembly is cotranslationally bound to the ER membrane and sorted to degradation.

Animals↗

PubMed1991

Evidence for the role of the secretory pattern of growth hormone in the regulation of serum concentrations of cholesterol and apolipoprotein E in rats.

Adult male Sprague-Dawley rats were hypophysectomized and connected to an automatic i.v. infusion system. The same daily dose of human GH (hGH) was given either as eight daily pulses (3-h intervals) to mimic the male specific secretory pattern of GH or as a continuous infusion of GH, to mimic the female secretory pattern. Hypophysectomized rats received i.v. replacement therapy with L-thyroxine and cortisol. The rats were treated for 5 days. The serum cholesterol concentration was higher when hGH was given continuously than when hGH was given as eight daily pulses. The concentration of high-density lipoprotein (HDL)-cholesterol was not influenced by intermittent GH treatment, but increased when hGH was given as a continuous infusion. The serum concentration of apolipoprotein (Apo) E increased following treatment with a continuous infusion of hGH, whereas eight daily pulses of hGH had no effect. The serum concentration of ApoA-I was unaffected by hGH treatment. The serum concentration of ApoB decreased to the same degree whether hGH was given as a continuous infusion or as eight daily pulses. The serum concentration of triglycerides was not affected by hGH treatment. These results indicate that the higher serum HDL-cholesterol and serum ApoE concentrations of female rats may be due to their more continuous secretion of GH. In contrast, the effects of GH on the serum concentration of ApoB, which is not sexually differentiated, may be independent of the mode of GH secretion.

Animals↗

PubMed1990

The assembly and secretion of apoB 100 containing lipoproteins in Hep G2 cells. Evidence for different sites for protein synthesis and lipoprotein assembly.

Pulse-chase studies combined with subcellular fractionation indicated that LpB 100 (i.e. the apoprotein B (apoB) 100 containing lipoproteins) was released to the lumen of the secretory pathway in a subcellular fraction enriched in smooth vesicles, and referred to as SMF (the smooth membrane fraction). The migration of SMF during gradient ultracentrifugation as well as kinetic studies indicated that the fraction was derived from a pre-Golgi compartment, probably the smooth endoplasmic reticulum (ER). Only small amounts of LpB 100 could be detected during these pulse-chase experiments in the subcellular fractions derived from the rough endoplasmatic reticulum (RER). SMF contained the major amount of the diacylglycerol acyltransferase activity present in the ER, while the major amount of membrane bound apoB 100 was present in the RER. Pulse-chase studies of the intracellular transfer of apoB 100 demonstrated the formation of a large membrane-bound preassembly pool in the ER, while no significant amount of apoB 100 radioactivity was present in the membrane of the Golgi apparatus. The maximal radioactivity of LpB 100, recovered from the ER or the Golgi lumen, was small compared with the radioactivity recovered from the ER membrane, indicating that the assembled LpB 100 rapidly leaves the cells. This in turn indicates that the rate-limiting step in the secretion of apoB 100 was the transfer of the protein from the ER membrane to the LpB 100 in the lumen. A portion of the intracellular pool of apoB 100 was not secreted but underwent posttranslational degradation.

Acetylglucosaminidase↗

PubMed1990

Apolipoprotein(a) and ischaemic heart disease in familial hypercholesterolaemia.

Serum concentrations of apolipoprotein(a) were measured in patients with heterozygous familial hypercholesterolaemia. The levels in 47 patients were a median of 2.5 times higher than those in controls matched for age and sex (240 [range 25-1245] vs 97 [7-1040] mg/l). Among patients with familial hypercholesterolaemia apo(a) levels were higher in those with (n = 48) than in those without (n = 72) ischaemic heart disease (283 [18-1245] vs 144 [7-741] mg/l); both in univariate and multivariate analysis serum apo(a) was the most significant variable distinguishing between the groups. Despite reducing LDL cholesterol by 30%, treatment with cholestyramine or pravastatin did not reduce apo(a) levels in these patients. These findings support the concept that apo(a) concentration is a genetic trait predisposing to ischaemic heart disease and imply that it may be useful in the identification of familial hypercholesterolaemia patients at high risk of coronary disease.

Adult↗

PubMed1990

Effect of short-term beta blockade on serum lipid levels and on the interaction of LDL with human arterial proteoglycans.

In view of conflicting evidence suggesting that beta-blockers have an anti-atherogenic effect as well as induce a potentially atherogenic lipoprotein profile, the effects of a short term beta-blockade on serum lipoproteins were studied in 39 healthy volunteers. Because the interaction of LDL with arterial proteoglycans appears to play a role in lipoprotein accumulation during atherogenesis, the effects of metoprolol and atenolol on low density lipoprotein interaction with human aortic proteoglycans were included in the study. We could confirm that the beta-blockers caused a decrease in HDL cholesterol and an increase in triglycerides, both potentially undesirable effects. In addition, however they induced a significant decrease in the in vitro LDL affinity for arterial proteoglycans. Since there appears to be a strong association between LDL reactivity with proteoglycans and risk for myocardial infarction, this effect of the beta-blockers may be an anti-atherogenic effect which overrides other effects on the lipoprotein pattern.

Adult↗

PubMed1990

Treatment of familial hypercholesterolaemia: a controlled trial of the effects of pravastatin or cholestyramine therapy on lipoprotein and apolipoprotein levels.

The efficacy and safety of a new, selective inhibitor of cholesterol synthesis, pravastatin, and the bile acid-binding resin, cholestyramine, were compared in a randomized, double-blind study of 120 patients with familial hypercholesterolaemia. After a run-in period of 8-10 weeks with assessment of dietary habits, the patients were treated with pravastatin + placebo, placebo + cholestyramine, or placebo alone. Active pravastatin therapy was initiated with 10 mg b.i.d. for 6 weeks, and was increased to 20 mg b.i.d. for the following 6 weeks. Cholestyramine was given at 24 g d-1, or the highest tolerable dose. After 6 weeks of therapy, serum total and LDL cholesterol levels were reduced by 17% and 21%, respectively, on pravastatin treatment, whereas the corresponding reductions with cholestyramine treatment were 24% and 30%, respectively. With an increased dose of pravastatin, serum and LDL cholesterol concentrations were reduced by 23% and 28%, respectively, after 12 weeks; the effect of cholestyramine was unchanged. HDL cholesterol levels increased in response to pravastatin, by 7% and 9% after 6 and 12 weeks, respectively. Concomitant changes in the concentrations of apolipoproteins B and AI were observed. Three patients discontinued the study because of side-effects: two subjects were treated with pravastatin and one was given placebo. The prevalence of side-effects (including laboratory abnormalities) was 35% for pravastatin, 30% for placebo, and 53% (significantly higher) for cholestyramine. We conclude that pravastatin, in a 40 mg daily dose, is as effective as cholestyramine in lowering LDL cholesterol in familial hypercholesterolaemia. Since the frequency of side-effects is higher with cholestyramine, pravastatin offers a promising alternative for the therapy of this genetic disease.

Adult↗

PubMed1989

Apolipoprotein B in human aortic biopsies in relation to serum lipids and lipoproteins.

A total of 46 patients, aged 39-71 years (mean 57.7), were studied. Forty-eight percent of the patients were hyperlipidemic and 63% had earlier suffered a myocardial infarction. Biopsies from aorta were obtained during coronary bypass surgery. Apo B was extracted from the intima by incubation of the tissue in buffer, followed by collagenase digestion. Intimal apo B was quantified in an immunoradiometric assay. There were significant correlations between total or collagenase-extractable apo B and serum cholesterol (rs = 0.39, P less than 0.01), serum triglycerides (rs = 0.33, P less than 0.05), LDL cholesterol (rs = 0.33, P less than 0.05) and serum apo B (rs = 0.37, P less than 0.05). The correlations were strongest for the collagenase-extractable apo B, while no correlations were observed for the buffer-extractable intimal apo B. No significant correlations were found between intimal apo B and serum HDL, apo A-I, smoking habits, history of hypertension or sustained myocardial infarction. Follow-up data were available for 42 of the patients, with a mean follow-up period of 35.1 months. The patients were classified according to symptoms of angina pectoris at the time of follow-up. There were significantly lower levels of serum apo A-I in the patients with poorer clinical prognosis. In a linear multiple stepwise regression analysis, apo A-I and serum LDL were significantly and independently related to clinical prognosis (R2 = 0.31).

Adult↗

PubMed1989

XbaI restriction fragment length polymorphism of apolipoprotein B in Swedish myocardial infarction patients.

Apolipoprotein B (apoB) is a major importance to the metabolism of lipoproteins, and there is also evidence which suggests that apoB plays a central role in atherogenesis. In order to study whether there is a link between one of the mutations of the apoB gene and premature coronary heart disease, the frequency of the XbaI RFLP for the apoB gene was analysed in 52 male myocardial infarction patients. These were compared with a control group matched for age and sex (n = 52), and a random population sample of middle-aged men (n = 106). Two alleles were identified by the presence (X2) or the absence (X1) of an XbaI cleavage site. A somewhat higher frequency of the X2 allele was seen among the patients, however there was no significant difference between patients and controls regarding the genotypes or allele frequencies. This observation does not confirm one earlier report where a higher frequency of the X1 allele was seen in myocardial infarction patients. Differences between the studied populations or epidemiological designs of the studies might explain the diverging results. Further studies are evidently needed to fully resolve the relation between the XbaI RFLP and risk of atherosclerotic disease or lipoprotein metabolism.

Adult↗

PubMed1989

Differential effects of continuous versus intermittent administration of growth hormone to hypophysectomized female rats on serum lipoproteins and their apoproteins.

The effects of the plasma pattern of GH on serum and lipoprotein levels of total cholesterol, triglycerides, apolipoprotein A-I (apo A-I), apolipoprotein B 48/100 (apo B), and apolipoprotein E (apo E) were studied in hypophysectomized female Sprague-Dawley rats, which had been given replacement therapy with L-T4 and hydrocortisone. Bovine GH (1 mg/kg.day) was administered sc either continuously by means of osmotic minipumps or by two daily injections. Serum lipoproteins were separated by sequential ultracentrifugation into very low density lipoproteins [density (d) less than 1.006 g/ml], low density lipoproteins (LDL; d 1.006-1.063 g/ml) and high density lipoproteins (HDL; d 1.063-1.21 g/ml). The content of total cholesterol and triglycerides were then determined. Apo A-I, apo B, and apo E were isolated from rat serum and antibodies raised in rabbits. In serum and in lipoprotein fractions, the content of apo A-I, apo-B, and apo E were determined by electroimmunoassay. After hypophysectomy, there occurred a decrease in serum cholesterol and serum levels of apo A-I and apo E, in spite of replacement therapy with T4 and cortisone. Similar changes were also observed in HDL. In contrast, apo B, cholesterol, and triglycerides were increased in LDL. Estradiol treatment had no effect on these changes. Continuous infusion of GH resulted in an increase in cholesterol and apo E in serum and HDL to the levels of intact females. In contrast, GH given twice daily had no effect. Therefore, the sexually dimorphic secretion of GH may be important for the regulation of sex differences in apo E and HDL cholesterol levels. There were no consistent effects of GH treatment on the levels of apo A-I in serum or HDL, but GH treatment resulted in a decrease in apo B and triglycerides in both serum and LDL, regardless of the mode of administration. This suggests that GH regulates the serum and LDL levels of apo B and triglycerides independently of the secretory pattern.

Animals↗

PubMed1988

Studies on the assembly of apo B-100-containing lipoproteins in HepG2 cells.

The relationship between apoB-100 and the membrane of the endoplasmic reticulum (ER) has been studied by a combination of pulse-chase methodology and subcellular fractionation. HepG2 cells were pulse-labeled with [35S]methionine for 3 min and chased with cold methionine for periods between 0 and 20 min. ApoB-100 and albumin, present in the membrane as well as in the luminal content of the ER vesicles, were isolated after each chase period. The results indicated that apoB-100 was cotranslationally bound to the membrane of the ER, and from this membrane-bound form, was transferred to the lumen after a delay of 10-15 min. Albumin was, as could be expected for a typical secretory protein, cotranslationally sequestered in the lumen of the ER. Apo-B-100-containing lipoproteins present in the microsomal lumen were analyzed by ultracentrifugation in a sucrose gradient. ApoB-100 occurred on rounded particles in three density regions: (i) d 1.1065-1.170 g/ml (Fraction I), (ii) d 1.011-1.045 g/ml (Fraction II), and (iii) d less than 1.011 g/ml (Fraction III). Fraction I, isolated from cells cultured in the absence of oleic acid, contained a homogenous population of particles with a mean diameter of approximately 200 A. Fraction I isolated from cells cultured in the presence of oleic acid was slightly more heterogeneous and had a mean diameter of approximately 250 A. Fractions II and III had mean diameters of 300 and 500 A, respectively. Cholesterol esters and triacylglycerol were the quantitatively dominating lipid constituents of all three fractions. Pulse-chase experiments indicated that Fraction I contained the newly assembled lipoproteins. With increasing chase time, the apoB-100 radioactivity was redistributed from Fraction I to Fractions II and III, indicating that Fraction I is converted into Fractions II and III during the intracellular transfer. Particles corresponding to Fractions II and III were by far the most abundant lipoproteins found in the medium. The results presented support the possibility of a sequential assembly of apoB-100-containing lipoproteins.

Albumins↗

PubMed1988

Aortic intimal lipid content and serum lipoproteins in patients undergoing coronary by-pass surgery as related to clinical prognosis.

Intimal lipid concentrations were determined in aortic biopsies obtained during coronary by-pass surgery. In addition serum lipoprotein and apolipoprotein levels were quantitated and their relationships to aortic intimal lipid concentrations were analysed. The possibility to use aortic intimal lipid and serum lipoprotein or apolipoprotein concentrations to predict clinical prognosis following the coronary by-pass operation was also evaluated. Intimal cholesterol, cholesterol ester, phosphatidylcholine and sphingomyelin were intercorrelated, whereas none of these lipid fractions correlated to aortic intimal triglyceride levels. Patients with hypertension had higher aortic intimal cholesterol ester levels than normotensive patients. There was a positive correlation between the number of stenosed coronary arteries and serum apo B or triglyceride levels. In addition there was a negative correlation between the number of stenosed arteries and HDL-cholesterol. Prognosis after the operation was inversely correlated to serum apo A-I levels. Our data do not, however, support the notion that aortic intimal lipid levels can be used to evaluate prognosis after coronary by-pass surgery.

Adult↗

PubMed1988

Quantification of plasma lipids and apolipoproteins in British Halflop rabbits. A comparison between normocholesterolemic rabbits, hypercholesterolemic rabbits (modified WHHL rabbits) and rabbits fed an atherogenic diet.

We have established isolation methods and developed electroimmunoassays for rabbit apolipoprotein A-I (apo A-I), apo B, apo C-III and apo E. The assays were used to characterize a hyperlipidemic strain of the British Halflop rabbits (BHL rabbits), obtained after cross-breeding with WHHL rabbits and referred to as modified WHHL rabbits, and to investigate the changes in the apolipoprotein levels induced by feeding normal BHL rabbits an atherogenic diet (0.25% cholesterol and 3% coconut oil). The modified WHHL rabbits were characterized by increased levels of apo B, apo C-III and apo E as well as cholesterol, phospholipids and triacylglycerol as compared to chow-fed BHL rabbits, while the apo A-I levels were only half of those found in the chow-fed animals. The modified WHHL rabbits had virtually no low density lipoprotein (LDL) receptor activity and a low fractional catabolic rate (FCR) of LDL. These results indicate that the modified WHHL rabbit has the homozygous form of the LDL receptor deficiency. The BHL rabbits fed the atherogenic diet showed increased levels of cholesterol, triacylglycerol, apo B, apo C-III and apo E, as compared to those of the chow-fed BHL rabbits. The apo E and apo C-III reached levels in the range of or even higher than those of the modified WHHL rabbits. The apo A-I levels on the other hand did not differ from those of the chow-fed rabbits. Feeding an atherogenic diet led to a decrease in the FCR of LDL to a level similar to that found in the modified WHHL rabbits.

Animals↗

PubMed1987

Structure and biosynthesis of apolipoprotein B.

Apolipoprotein (apo) B100 copy deoxyribonucleic acid (cDNA) has been cloned and sequenced. The total sequence of apo B100 messenger ribonucleic acid has been revealed by the work from our group, as well as other groups. The sequence spans 13,689 nucleotides from the initiation to the stop codon. Thus the messenger has the capacity to code for 4563 amino acids corresponding to a molecular weight of approximately 510,000. Computer analyses revealed the presence of regions with amphipathic alpha-helix and of hydrophobic regions with a high probability of beta-structure. Regions of the molecule are characterized by continuous variations between hydrophillic and hydrophobic sequences, the latter coinciding with a high probability of beta-structure. It is suggested that this beta-structure is involved, together with the amphipathic alpha-helix, in the binding of apo B100 to the lipid. Pulse-chase studies in Hep G2 cells showed that apo B100 is synthesized as one protein, with a translation time of 14 minutes. The protein is transferred through the cell and secreted within 30 minutes without undergoing any major change in molecular mass. The residence kinetics in the endoplasmic reticulum (ER) is characterized by an increase during the first 10 to 15 minutes of chase followed by an almost linear decrease with a decay rate of 6%/min. The transfer through the ER to the Golgi apparatus accounts for one third of the time needed for the intracellular transfer of apo B100, whereas two thirds of the time is required for the transfer through the later part of the secretory pathway.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗

PubMed1987

Apolipoprotein B: structure, biosynthesis and role in the lipoprotein assembly process.

The complete amino acid sequence of the liver-synthesized apolipoprotein B (apoB) species, apoB 100, has been derived from cloned cDNA. The protein consists of 4536 amino acids (+ a 27 amino acid signal sequence). Cysteine is clustered in the N-terminal 1/10 of the protein, suggesting the presence of a stabilized tertiary structure in this part of the molecule. Three types of structure are suggested to be of importance for the binding of the protein to lipids; (i) hydrophobic sequences with a high probability for beta-sheet structure, (ii) strict amphipathic beta-sheets, and (iii) amphipathic alfa-helices. An apoB 100 molecule is completed within 10-14 min and secreted after approximately 30 min, 1/3 of which is due to the transfer through the endoplasmic reticulum (ER), while 2/3 is spent in the Golgi apparatus. ApoB 100 is co-translationally N-glycosylated and 25% of the oligosaccharide chains is processed in the Golgi compartment. Other posttranslational modifications that have been discussed include covalent acylation and phosphorylation. It has also been suggested that the lipid moiety of the apoB 100 lipoproteins are modified during the passage through the Golgi apparatus. The site of lipoprotein assembly is suggested to be separated from the site of apoB 100 synthesis, and apoB 100 appears to be co-translationally bound to the ER membrane and from this transferred to the ER lumen. Based on these observations a model for the assembly of apoB 100 lipoproteins is discussed in this paper. The intestinal derived apoB species, apoB 48, has a molecular mass of 210 kDa and appears to correspond to the N-terminal 48% of apoB 100. The mechanism by which apoB 48 is formed is still not known. Available data indicate that the protein is formed within the intestinal cells, these data also argue against the possibility that apoB 48 is formed by posttranslational proteolysis of apoB 100. The formation of a separate apoB 48 mRNA by alternative splicing has been suggested, based on the observation of a 7 kb mRNA which corresponds to the 5' portion of the apoB 100 mRNA. However, the most abundant apoB mRNA species found in the intestine have a size that corresponds to that of the apoB 100 mRNA, furthermore the observation that apoB 48 appears to terminate in a 7.5 kb exon that appears to lack alternative splice sites, does not favour the possibility of alternative splicing.

Amino Acid Sequence↗