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Biomedical subjects

C Bruce

Publications and source records attributed to C Bruce.

At least 37 records · Page 2Linked to original sources
PubMed1999

Observational study of maternal anthropometry and fetal insulin.

AIMS: To examine the relation between maternal body fat and fetal metabolism. METHODS: In this observational study, cord blood samples were collected from 60 infants of healthy women for the measurement of insulin and C peptide concentrations. Maternal weight, height, body mass index (BMI) and body composition (skinfold thickness measurements and bioelectrical impedance) were assessed at 13-15 weeks of gestation. Twenty five of the volunteers agreed to have a 75 g oral glucose tolerance test at 28-31 weeks of gestation. RESULTS: Positive correlations were observed with both cord insulin or C peptide concentrations and maternal early pregnancy BMI (r=0.44, p=0.002 and r=0.33, p=0.008, respectively). There was no significant correlation between cord insulin or C peptide concentrations and birthweight or birth weight centiles. CONCLUSION: Maternal BMI could be a predictor of fetal cord insulin concentration.

Anthropometry↗
PubMed1999

Increased atherosclerosis in ApoE and LDL receptor gene knock-out mice as a result of human cholesteryl ester transfer protein transgene expression.

The plasma cholesteryl ester transfer protein (CETP) plays a major role in the catabolism of HDL cholesteryl ester (CE). CETP transgenic mice have decreased HDL cholesterol levels and have been reported to have either increased or decreased early atherosclerotic lesions. To evaluate the impact of CETP expression on more advanced forms of atherosclerosis, we have cross-bred the human CETP transgene into the apoE knock-out (apoE0) background with and without concomitant expression of the human apo A-I transgene. In this model the CETP transgene is induced to produce plasma CETP levels 5 to 10 times normal human levels. CETP expression resulted in moderately reduced HDL cholesterol (34%) in apoE0 mice and markedly reduced HDL cholesterol (76%) in apoE0/apoA1 transgenic mice. After injection of radiolabeled HDL CE, the CETP transgene significantly delayed the clearance of CE radioactivity from plasma in apoE0 mice, but accelerated the clearance in apoE0/apoA1 transgenic mice. ApoE0/CETP mice displayed an increase in mean atherosclerotic lesion area on the chow diet (approximately 2-fold after 2 to 4 months, and 1.4- to 1.6-fold after 7 months) compared with apoE0 mice (P<0.02). At 7 months apoA1 transgene expression resulted in a 3-fold reduction in mean lesion area in apoE0 mice (P<0.001). In the apoE0/apoA1 background, CETP produced an insignificant 1.3- to 1.7-fold increase in lesion area. In further studies the CETP transgene was bred onto the LDL receptor knock-out background (LDLR0). After 3 months on the Western diet, the mean lesion area was increased 1.8-fold (P<0.01) in LDLR0/CETP mice, compared with LDLR0 mice. These studies indicate that CETP expression leads to a moderate increase in atherosclerosis in apoE0 and LDLR0 mice, and suggest a proatherogenic effect of CETP activity in metabolic settings in which clearance of remnants or LDL is severely impaired. However, apoA1 overexpression has more dramatic protective effects on atherosclerosis in apoE0 mice, which are not significantly reversed by concomitant expression of CETP.

Animals↗
PubMed1999

Targeted mutation of plasma phospholipid transfer protein gene markedly reduces high-density lipoprotein levels.

It has been proposed that the plasma phospholipid transfer protein (PLTP) facilitates the transfer of phospholipids and cholesterol from triglyceride-rich lipoproteins (TRL) into high-density lipoproteins (HDL). To evaluate the in vivo role of PLTP in lipoprotein metabolism, we used homologous recombination in embryonic stem cells and produced mice with no PLTP gene expression. Analysis of plasma of F2 homozygous PLTP-/- mice showed complete loss of phosphatidylcholine, phosphatidylethanolamine, phosphatidylinositol, sphingomyelin, and partial loss of free cholesterol transfer activities. Moreover, the in vivo transfer of [3H]phosphatidylcholine ether from very-low-density proteins (VLDL) to HDL was abolished in PLTP-/- mice. On a chow diet, PLTP-/- mice showed marked decreases in HDL phospholipid (60%), cholesterol (65%), and apo AI (85%), but no significant change in non-HDL lipid or apo B levels, compared with wild-type littermates. On a high-fat diet, HDL levels were similarly decreased, but there was also an increase in VLDL and LDL phospholipids (210%), free cholesterol (60%), and cholesteryl ester (40%) without change in apo B levels, suggesting accumulation of surface components of TRL. Vesicular lipoproteins were shown by negative-stain electron microscopy of the free cholesterol- and phospholipid-enriched IDL/LDL fraction. Thus, PLTP is the major factor facilitating transfer of VLDL phospholipid into HDL. Reduced plasma PLTP activity causes markedly decreased HDL lipid and apoprotein, demonstrating the importance of transfer of surface components of TRL in the maintenance of HDL levels. Vesicular lipoproteins accumulating in PLTP-/- mice on a high-fat diet could influence the development of atherosclerosis.

Animals↗
PubMed1999

Lateral shelf acetabuloplasty in Perthes' disease. A review of the end of growth.

The surgical treatment of Perthes' disease by femoral or innominate osteotomy is not as effective in those over the age of eight years as it is in the younger child. This has prompted the search for other types of management in those who are older. The preliminary results of the use of a lateral shelf acetabuloplasty for such cases have shown encouraging results at two years. The concern with such an operation is that it might interfere with the growth of the outer aspect of the acetabulum and so prejudice the long-term outcome. We describe a review at maturity of 26 children presenting with early disease after the age of eight years who were treated by lateral shelf acetabuloplasty. The results suggest that the outcome is improved; 22 of 27 hips were rated as Stulberg groups 1 to 3. Poor results occurred in children, particularly girls, presenting with Group-4 disease over the age of 11 years.

Acetabulum↗
PubMed1998

Radio-enzymatic determination of histamine: interference by fluoride and possible activation of histamine methyl transferase.

The reproducibility of a radio-enzymatic method for determining plasma histamine was found to be affected by the anti-coagulant used for collecting blood. Recovery experiments from whole blood indicated that heparin yielded values that were more accurate than with EDTA or sodium fluoride; fluoride gave a mean value which was +41% higher than with heparin (P=0.054). Addition of fluoride to a standard calibration curve, and of increasing amounts to aliquots of 5 ng histamine also yielded higher values than in controls, up to +15% (P<0.1) and +14.1% (P=0.03) respectively. Fluoride did not affect the detecting system and was not contaminated with histamine; nor did it breakdown the methyl donor, S-adenosyl-L-methionine. It is concluded that heparin is the anti-coagulant of choice and that fluoride may activate histamine methyl transferase from pig brain. Fluoride may possibly have a biological role as an enzyme-activator and a usefulness in the therapy of mastocytosis.

Edetic Acid↗
PubMed1998

The implications of the structure of the bactericidal/permeability-increasing protein on the lipid-transfer function of the cholesteryl ester transfer protein.

The cholesteryl ester transfer protein (CETP) is evolutionarily related to the bactericidal/permeability-increasing protein (BPI). The recently solved structure of BPI shows an elongated, boomerang-shaped molecule, with two hydrophobic pockets opening to its concave side. These pockets each contain a phospholipid molecule. A model of CETP, based on the recently solved crystal structure of BPI, provides the basis for interpreting functional studies on CETP. In this model, C-terminal residues 461-476, which were shown to be required for neutral lipid transfer between plasma lipoproteins, from an amphipathic helix covering the opening of the N-terminal pocket. A possible lipid-transfer mechanism for CETP, with the initial step involving the disordering of lipids in the lipoprotein surface, followed by the flipping and entry of a lipid molecule into the hydrophobic lipid-binding pocket, is hypothesized in light of structural evidence and recent studies.

Antimicrobial Cationic Peptides↗
PubMed1998

Physiological changes in insulin resistance in human pregnancy: longitudinal study with the hyperinsulinaemic euglycaemic clamp technique.

OBJECTIVE: To perform measurements of insulin resistance serially in pregnancy and after childbirth in normal women. DESIGN: Longitudinal study. SETTING: Teaching hospital. METHODS: Ten normal women were studied using the hyperinsulinaemic euglycaemic clamp technique before pregnancy, at 16, 26 and 36 weeks gestation, and 8 weeks after delivery. All women breastfed their infants. Insulin resistance was measured by the glucose infusion rate required to maintain the plasma glucose at 4.5 mmol/l. RESULTS: There was a progressive increase in insulin resistance in all women as pregnancy progressed. The increase in resistance was most marked between 16 and 26 weeks of gestation, with only minimal progression thereafter. Lactation does not alter insulin resistance. CONCLUSION: The timing and extent of changes in insulin resistance are of physiological importance because they affect all classes of maternal and fetal substrates.

Adult↗
PubMed1998

Plasma lipid transfer proteins, high-density lipoproteins, and reverse cholesterol transport.

Cholesteryl ester transfer protein (CETP) and phospholipid transfer protein (PLTP) are members of the lipid transfer/lipopolysaccharide binding protein gene family. Recently, the crystal structure of one of the members of the gene family, bactericidal permeability increasing protein, was solved, providing potential insights into the mechanisms of action of CETP and PLTP. These molecules contain intrinsic lipid binding sites and appear to act as carrier proteins that shuttle between lipoproteins to redistribute lipids. The phenotype of human CETP genetic deficiency states and CETP transgenic mice indicates that CETP plays a major role in the catabolism of high-density lipoprotein (HDL) cholesteryl esters and thereby influences the concentration, apolipoprotein content, and size of HDL particles in plasma. PLTP also appears to have an important role in determining HDL levels and speciation. Recent data indicate that genetic CETP deficiency is associates with an excess of coronary heart disease in humans, despite increased HDL levels. Also, CETP expression is anti-atherogenic in many mouse models, even while lowering HDL. These data tend to support the reverse cholesterol transport hypothesis, i.e., that anti-atherogenic properties of HDL are related to its role in reverse cholesterol transport. Recently, another key molecule involved in this pathway was identified, scavenger receptor BI; this mediates the selective uptake of HDL cholesteryl esters in the liver and thus constitutes a pathway of reverse cholesterol transport parallel to that mediated by CETP. Reflecting its role in reverse cholesterol transport, the CETP gene is up-regulated in peripheral tissues and liver in responses to dietary or endogenous hypercholesterolemia. An analysis of the CETP proximal promoter indicates that it contains sterol regulatory elements highly homologous to those present in 3-hydroxy-3-methylglutaryl-coenzyme A reductase; the CETP gene is transactivated by the binding of SREBP-1 to these elements. A challenge for the future will be the manipulation of components of the reverse cholesterol transport pathway, such as CETP, PLTP, or scavenger receptor BI for therapeutic benefit.

Animals↗
PubMed1998

Relationship of HDL and coronary heart disease to a common amino acid polymorphism in the cholesteryl ester transfer protein in men with and without hypertriglyceridemia.

Plasma triglyceride (TG) levels are inversely related to HDL-cholesterol levels and subjects with high TG and low HDL cholesterol have increased coronary heart disease (CHD) risk. Plasma cholesteryl ester transfer protein (CETP) transfers cholesteryl esters from HDL to TG-rich lipoproteins. In this study we determined the relationship between a common CETP amino acid polymorphism (I405V) and CETP and HDL levels and CHD prevalence in 576 men of Japanese ancestry in the Honolulu Heart Program cohort. This conservative substitution was associated with altered plasma CETP concentration (1.95 +/- 0.54, 1.91 +/- 0.57, and 1.77 +/- 0.57 microg/ml for the II, IV and VV genotypes, respectively). The distribution of plasma CETP concentrations among the VV, but not II, men appeared bimodal (P < 0.01), suggesting the presence of a functionally significant CETP gene mutation(s) in a subset of V alleles. HDL-C levels were higher in VV than IV for II men (55.4 +/- 17.4, 51.3 +/- 16.6, 51.1 +/- 17.0 mg/dl, P < 0.04). However, the increase in HDL was only significant in VV men with plasma TG > 165 mg/dl. Although CHD prevalence was not significantly different among the three genotypes in this population, in the subpopulation with high plasma TG, CHD prevalence appeared higher among VV than IV or II subjects (38% vs. 27% vs. 18%, P < 0.05 for an interaction of genotype and plasma TG levels). In fresh plasma from a separate group of normolipidemic subjects, the V/I polymorphism was not associated with any change in plasma CETP specific activity. The data suggest that a widespread and common CETP gene mutation(s) in linkage disequilibrium with 405V causes low CETP. Among hypertriglyceridemic men this is associated with higher HDL and possibly with increased CHD.

Amino Acids↗
PubMed1998

Complement factor H or a related protein is a marker for transitional cell cancer of the bladder.

The BTAstat and BTA TRAK tests are new immunoassays that detect and measure an antigen in the urine of individuals diagnosed with bladder cancer. As described in this report, the monoclonal antibodies used in these kits were developed by immunizing mice with partially purified protein preparations derived from the urine of patients with bladder cancer. The antigen that is recognized by the monoclonal antibodies was purified from the urine of bladder cancer patients by immunoaffinity chromatography and identified as being either complement factor H (FH) or a closely related protein (CFHrp) by partial amino acid sequence analysis. Like serum FH, the urine antigen was demonstrated to have a complement factor C3b binding site and to accelerate the degradation of C3b in the presence of complement factor I. The culture supernatants from several human bladder, cervical, and renal cancer cell lines contained antigen as determined by immunoassay, and antigen affinity-purified from HeLaS3 culture media was shown to have FH activity. Moreover, the cell lines were shown to make products of the expected sizes by reverse transcription-PCR using FH-specific primers. In contrast, normal human epithelial keratinocytes, a myeloid leukemia cell line, and the colon cancer line LS174T were negative for production of a FH-like protein (CFHrp). We propose that the expression of proteins with FH-like activities may confer a selective growth advantage to cancer cells in vivo by decreasing complement activity, thus aiding their escape from lysis by immune surveillance. Identification of these proteins as cancer products also suggests avenues of chemotherapy or immunotherapy of some cancers.

Amino Acid Sequence↗
PubMed1997

Involvement of tyrosine phosphorylation in endothelin-1-induced calcium-sensitization in rat small mesenteric arteries.

1. We have studied the effect of endothelin-1 stimulation on protein tyrosine phosphorylation levels in intact small mesenteric arteries of the rat and investigated the effects of tyrosine kinase inhibition on the contractile response to this agonist. 2. Endothelin-1 stimulated a rapid (20 s), sustained (up to 20 min) and concentration-dependent (1-100 nM) increase in protein tyrosine phosphorylation levels which coincided temporally with the contractile response in intact and alpha-toxin permeabilized small artery preparations. Tyrosine phosphorylation was increased in four main clusters of proteins of apparent molecular mass 28-33, 56-61, 75-85 and 105-115 kDa. Endothelin-1-induced protein tyrosine phosphorylation was independent of extracellular calcium, antagonized by the tyrosine kinase inhibitor tyrphostin A23 but not by the inactive tyrphostin A1. 3. In intact small arteries tyrphostin A23 inhibited the force developed to endothelin-1 at all concentrations studied; at higher concentrations (10 and 100 nM) the profile of contraction was altered from a sustained to a transient response. Tyrphostin A1 inhibited the contractile response to endothelin-1 at all concentrations except 100 nM; the profile of the response was not altered. Neither tyrphostin affected the transient phasic contraction induced by endothelin-1 (100 nM) in the absence of extracellular calcium. 4. In rat alpha-toxin permeabilized mesenteric arteries endothelin-1 caused a concentration-dependent increase in force in the presence of 10 microM GTP and low (pCa 6.7) constant calcium, demonstrating increased sensitivity of the contractile apparatus to calcium. Tyrphostin A23 inhibited this response by approximately 50%, tyrphostin A1 did not affect endothelin-1-induced calcium sensitization of force. 5. We conclude that increased tyrosine phosphorylation is important in the contractile response induced by endothelin-1 in intact small mesenteric arteries. Furthermore our data implicate activation of this signalling pathway in the tonic phase of contraction possibly through modulation of the sensitivity of the contractile apparatus to calcium.

Animals↗
PubMed1996

Human cholesteryl ester transfer protein gene proximal promoter contains dietary cholesterol positive responsive elements and mediates expression in small intestine and periphery while predominant liver and spleen expression is controlled by 5'-distal sequences. Cis-acting sequences mapped in transgenic mice.

The plasma cholesteryl ester transfer protein (CETP) facilitates the transfer of high density lipoprotein cholesteryl esters to other lipoproteins and appears to be a key regulated component of reverse cholesterol transport. Earlier studies showed that a CETP transgene containing natural flanking sequences (-3.4 kilobase pairs (kbp) upstream, +2.2 kbp downstream) was expressed in an authentic tissue distribution and induced in liver and other tissues in response to dietary or endogenous hypercholesterolemia. In order to localize the DNA elements responsible for these effects, we prepared transgenic mice expressing six new DNA constructs containing different amounts of natural flanking sequence of the CETP gene. Tissue-specific expression and dietary cholesterol response of CETP mRNA were determined. The native pattern of predominant expression in liver and spleen with cholesterol induction was shown by a -3.4 (5'), +0.2 (3') kbp transgene, indicating no major contribution of distal 3'-sequences. Serial 5'-deletions showed that a -570 base pairs (bp) transgene gave predominant expression in small intestine with cholesterol induction of CETP mRNA in that organ, and a -370 bp transgene gave highest expression in adrenal gland with partial dietary cholesterol induction of CETP mRNA and plasma activity. Further deletion to -138 bp 5'-flanking sequence resulted in a transgene that was not expressed in vivo. Both the -3.4 kbp and -138 bp transgenes were expressed when transfected into a cultured murine hepatocyte cell line, but only the former was induced by treating the cells with LDL. When linked to a human apoA-I transgene, the -570 to -138 segment of the CETP gene promoter gave rise to a relative positive response of hepatic apoA-I mRNA to the high cholesterol diet in two out of three transgenic lines. Thus, 5'-elements between -3,400 and -570 bp in the CETP promoter endow predominant expression in liver and spleen. Elements between -570 and -370 are required for expression in small intestine and some other tissues, and elements between -370 and -138 contribute to adrenal expression. The minimal CETP promoter element associated with a positive sterol response in vivo was found in the proximal CETP gene promoter between -370 and -138 bp. This region contains a tandem repeat of a sequence known to mediate sterol down-regulation of the HMG-CoA reductase gene, suggesting either the presence of separate positive and negative sterol response elements in this region or the use of a common DNA element for both positive and negative sterol responses.

Animals↗
PubMed1996

Increased prebeta-high density lipoprotein, apolipoprotein AI, and phospholipid in mice expressing the human phospholipid transfer protein and human apolipoprotein AI transgenes.

Human plasma phospholipid transfer protein (PLTP) circulates bound to high density lipoprotein (HDL) and mediates both net transfer and exchange of phospholipids between different lipoproteins. However, its overall function in lipoprotein metabolism is unknown. To assess the effects of increased plasma levels of PLTP, human PLTP transgenic mice were established using the human PLTP gene driven by its natural promoter. One line of PLTP transgenic mice with moderate expression of PLTP mRNA and protein was obtained. The order of human PLTP mRNA expression in tissues was: liver, kidney, brain, small intestine > lung > spleen > heart, adipose tissue. Western blotting using a human PLTP monoclonal antibody revealed authentic human PLTP (Mr 80 kD) in plasma. Plasma PLTP activity was increased by 29% in PLTP transgenic mice. However, plasma lipoprotein analysis, comparing PLTP transgenic mice to control littermates, revealed no significant changes in the plasma lipoprotein lipids or apolipoproteins. Since previous studies have shown that human cholesteryl ester transfer protein and lecithin:cholesterol acyltransferase only function optimally in human apoAI transgenic mice, the human PLTP transgenic mice were cross-bred with human apoAI transgenic mice. In the human apoAI transgenic background, PLTP expression resulted in increased PLTP activity (47%), HDL phospholipid (26%), cholesteryl ester (24%), free cholesterol (37%), and apoAI (22%). There was a major increase of apoAI in prebeta-HDL (56%) and a small increase in alpha-HDL (14%). The size distribution of HDL particles within alpha- and prebeta-migrating species was not changed. The results suggest that PLTP increases the influx of phospholipid and secondarily cholesterol into HDL, leading to an increase in potentially antiatherogenic prebeta-HDL particles.

Animals↗
PubMed1996

Increased coronary heart disease in Japanese-American men with mutation in the cholesteryl ester transfer protein gene despite increased HDL levels.

Plasma high density lipoprotein (HDL) levels are strongly genetically determined and show a general inverse relationship with coronary heart disease (CHD). The cholesteryl ester transfer protein (CETP) mediates the transfer of cholesteryl esters from HDL to other lipoproteins and is a key participant in the reverse transport of cholesterol from the periphery to the liver. A high prevalence of two different CETP gene mutations (D442G, 5.1%; intron 14G:A, 0.5%), was found in 3,469 men of Japanese ancestry in the Honolulu Heart Program and mutations were associated with decreased CETP (-35%) and increased HDL chol levels (+10% for D442G). However, the overall prevalence of definite CHD was 21% in men with mutations and 16% in men without mutations. The relative risk (RR) of CHD was 1.43 in men with mutations (P < .05); after adjustment for CHD risk factors, the RR was 1.55 (P = .02); after additional adjustment for HDL levels, the RR was 1.68 (P = .008). Similar RR values were obtained for the D442G mutation alone. Increased CHD in men with mutations was primarily observed for HDL chol 41-60 mg/dl; for HDL chol > 60 mg/dl men with and without mutations had low CHD prevalence. Thus, genetic CETP deficiency appears to be an independent risk factor for CHD, primarily due to increased CHD prevalence in men with the D442G mutation and HDL cholesterol between 41 and 60 mg/dl. The findings suggest that both HDL concentration and the dynamics of cholesterol transport through HDL (i.e., reverse cholesterol transport) determine the anti-atherogenicity of the HDL fraction.

Aged↗
PubMed1996

Plasma histamine in patients with tropical disorders and acute asthma who exhibit eosinophilia.

The mean venous plasma histamine of 14 patients with eosinophilia was 18.7 nmol/L (range 0-86.4) and significantly higher than that of 4.0 nmol/L for 29 normal subjects [range 0-10.8 (P < 0.01)]. The eosinophilia of schistosomiasis (two patients), strongyloidiasis (one patient), and asthma (four patients) was accompanied by raised plasma histamine concentrations. Normal levels occurred in onchocerciasis (four patients) and in two of three patients with non-specific eosinophilia. Plasma histamine concentrations in the tropical disorders are not thought to have been reported previously. The initial arterial and venous plasma histamine concentrations of 14 acute asthmatics, were significantly positively correlated with the respective arterial (P < 0.01) and venous (P < 0.05) eosinophil counts. The results in acute asthma support the hypothesis that increased plasma histamine may have a role in promoting eosinophil release from the bone-marrow. Eosinophilia is not dependent upon a raised plasma histamine in some disorders, but may be in others, such as schistosomiasis.

Acute Disease↗
PubMed1995

Regulation of murine plasma phospholipid transfer protein activity and mRNA levels by lipopolysaccharide and high cholesterol diet.

Plasma phospholipid transfer protein mediates the net movement of phospholipids between lipoproteins and between lipid bilayers and high density lipoprotein. In this study, the mouse phospholipid transfer protein cDNA was cloned by reverse transcription polymerase chain reactions based on the cDNA sequence of human phospholipid transfer protein. The predicted amino acid sequence of mouse phospholipid transfer protein shows the protein to be 476 amino acids long and to have a sequence identity of 83% with that of human phospholipid transfer protein. Mouse plasma phospholipid transfer protein activity is 1.5-2 times that of human plasma phospholipid transfer protein activity. As in humans, mouse peripheral tissues displayed a higher abundance of phospholipid transfer protein mRNA than observed in central organs. The order of phospholipid transfer protein mRNA expression was as follows: lung > adipose tissue, placenta, testis > brain > muscle, heart, liver. We examined the regulation of phospholipid transfer protein expression by dietary cholesterol and by bacterial lipopolysaccharide. A high fat, high cholesterol diet caused a significant increase (35%) in plasma phospholipid transfer protein activity and a significant increase (18%) in high density lipoprotein phospholipids. This increased activity was accompanied by approximately 100% increase in phospholipid transfer protein mRNA in lung. After lipopolysaccharide injection, plasma phospholipid transfer protein activity was decreased by approximately 66%. This decrease in activity was associated with a similar decrease in phospholipid transfer protein mRNA in lung, adipose tissue, and liver. The decrease in plasma phospholipid transfer protein activity was also associated with a significant increase (17%) in high density lipoprotein phospholipid concentration. The opposite changes in phospholipids levels with lipopolysaccharide treatment and dietary cholesterol despite similarly increased high density lipoprotein phospholipids levels indicate that high density lipoprotein phospholipids levels are likely determined both by phospholipid transfer protein levels and by gradients of phospholipids concentration between high density lipoprotein and other phospholipids sources.

Amino Acid Sequence↗
PubMed1995

Point mutagenesis of positively charged amino acids of cholesteryl ester transfer protein: conserved residues within the lipid transfer/lipopolysaccharide binding protein gene family essential for function.

The cholesteryl ester transfer protein (CETP) binds to plasma lipoproteins and transfers neutral lipids between them. Previous studies showed that lipoprotein binding involves ionic interactions between CETP and lipoproteins, with increased binding of CETP to lipoproteins carrying increased negative charge. In order to understand the molecular determinants of lipoprotein binding in CETP, site-directed mutagenesis was carried out on positively charged amino acids within and outside regions of conserved sequence in the putative family of lipid transfer/lipopolysaccharide (LPS) binding proteins (LT/LBP). Within the conserved regions, two mutant proteins, K233A and R259D, were well secreted by the transfected cells but showed markedly reduced cholesteryl ester transfer activity. Separating the bound from free CETP by gel filtration after incubation with HDL, HDL binding by K233A was found to be impaired, suggesting that the binding deficiency of the mutant may be responsible for decreased transfer activity. Kinetic analysis showed a marked increase in the apparent Km but no change in Vmax, consistent with a lipoprotein binding defect. Thus, within CETP, K233 and R259 play an essential role in cholesteryl ester transfer activity probably by mediating binding of CETP to lipoproteins. Sequence alignment of CETP, phospholipid transfer protein, LPS binding protein, and bactericidal permeability-inducing protein showed that K223 and R259 were strictly conserved as positively charged amino acids, suggesting a common function within the LT/LBP gene family.

Acute-Phase Proteins↗