Detection and isolation of recombinant proteins from mammalian cells by immunoaffinity chromatography: p53.
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Amino Acid Sequence↗
Biomedical subjects
B Sepehrnia
Publications and source records attributed to B Sepehrnia.
Heat shock protein 84 forms a complex with mutant p53 protein predominantly within a cytoplasmic compartment of the cell.
Cellular DNA damage results in the increased expression and accumulation of the p53 tumor suppressor protein within the nucleus which leads to cell cycle arrest or apoptosis. In some cases, however, wild-type p53 and some mutant forms of p53 reside in the cytoplasm of cancer cells. To understand the mechanism responsible for its cytoplasmic retention, studies were undertaken to determine if unique proteins form a complex with mutant p53 within the cytoplasm of transformed cells. One protein, with an apparent molecular mass of 92 kDa (p92), was observed to form a complex with a temperature-sensitive mutant p53 (TSp53(Val-135)) in the cytoplasm of transformed rat embryo fibroblasts at the non-permissive temperature. p92 copurified with TSp53(Val-135) on a p53-specific immunoaffinity column and a gel filtration column. The protein was purified to homogeneity and identified as hsp84 by partial amino acid sequence analysis. hsp84 is a member of the hsp90 class of proteins. At the non-permissive temperature, TSp53(Val-135) and hsp84 colocalized in the cytoplasm near the nuclear envelope. At the permissive temperature, TSp53(Val-135) resides in the nucleus and expresses a "wild-type like" conformation. Under these conditions hsp84 continued to reside in the cytoplasm and little or no hsp84 formed a complex with p53. The results suggest that hsp84 binds mutant p53 in a spatial and/or conformation dependent manner.
Amino Acid Sequence↗
Screening for mtDNA diabetes mutations in Pima Indians with NIDDM.
More than half of the Pima Indians over age 35 years have non-insulin-dependent (type II) diabetes mellitus (NIDDM). Extensive data indicate the importance of maternal diabetes in determining their risk for diabetes. Generally, the risk of having NIDDM is higher in patients with affected mothers than affected fathers. This has been attributed to intrauterine factors, but recently mitochondrial inheritance has been raised as an alternative hypothesis. In other populations, several families and individuals with diabetes due to a mitochondrial DNA point mutation at nucleotide 3243 in the tRNA(leu(UUR)) gene have been described, as has one family with a 10.4 kb mitochondrial DNA duplication/deletion. We tested whether these specific mitochondrial gene mutations could explain a portion of the excess maternal transmission seen in the Pima Indians. Mitochondrial DNA obtained from blood lymphocytes of 148 Pima Indians with NIDDM was screened both for the point mutation at nt 3243, and the 10.4 kb duplication/deletion. Neither of these mutations was detected, and although a small proportion of the excess maternal transmission in Pima Indians could still be due to yet undescribed mitochondrial mutations or imprinted nuclear genes, our data support the role of the intrauterine environment in this population.
Adolescent↗
Genetic studies of human apolipoproteins. XX. Genetic polymorphism of apolipoprotein J and its impact on quantitative lipid traits in normolipidemic subjects.
Apolipoprotein J (apo J) is a newly identified member of a growing family of proteins associated with various lipoprotein particles. Apo J is a glycoprotein which exists in the plasma associated with high-density lipoprotein subfractions which also contain apo A-I and cholesteryl ester transfer protein (CETP). We have investigated the possible existence of genetic polymorphism at the apo J structural locus and have evaluated its role in lipid metabolism. By employing isoelectric focusing and immunoblotting techniques, we have screened plasma or serum samples from six population groups: U.S. whites, Amerindians, Eskimos, New Guineans, U.S. blacks, and Nigerian blacks. Apo J revealed a common two-allele polymorphism only in populations with African ancestry and was found to be monomorphic in all other population groups tested. The genetic basis of the two alleles designated--APO J*1 and APO J*2, at a single structural locus, apo J-- was confirmed in a large number of segregating families. In the U.S. blacks, the frequencies of the APO J*1 and APO J*2 alleles were .76 and .24, respectively, and in the Nigerian blacks these values were .72 and .28, respectively. In addition, a single example of a rare allele designated APO J*3 was also encountered in the U.S. black sample. In Nigerian blacks, the apo J polymorphism's impact on seven quantitative lipid traits--total cholesterol, LDL-cholesterol, HDL-cholesterol, HDL3-cholesterol, HDL2-cholesterol, VLDL-cholesterol, and triglycerides--was investigated. No significant impact of the apo J polymorphism was observed for any of these lipid traits.
Adolescent↗
Highly polymorphic apolipoprotein A-IV locus in the baboon.
Apolipoprotein A-IV is found in mesenteric lymph chylomicrons, very low density lipoprotein particles, high density lipoprotein particles, and in the lipoprotein-free fraction of plasma. Apolipoprotein A-IV is polymorphic in a variety of species including human, dog, and horse. Efforts to estimate the impact of apolipoprotein A-IV structural variation on quantitative lipid levels in humans have been limited by the low frequency of the less common alleles. In the baboon, Papio hamadryas anubis, we have found apolipoprotein A-IV to be highly variable at the protein level with five alleles appearing at polymorphic frequency. We have confirmed the autosomal codominant inheritance of these five alleles in pedigreed baboons. The baboon has been shown to be a suitable animal model for the study of atherosclerosis, and the existence of a common, multi-allele apolipoprotein A-IV polymorphism in the baboon may be useful in elucidating the role of apolipoprotein A-IV in lipid metabolism.
Alleles↗
Genetic studies of human apolipoproteins. VIII. Role of the apolipoprotein H polymorphism in relation to serum lipoprotein concentrations.
Apolipoprotein H (APO H) has recently been identified as a structural component of chylomicrons, very low-density lipoproteins (VLDL), low-density lipoproteins (LDL), and high-density lipoproteins (HDL). Although the precise metabolic function of APO H in lipid metabolism is not certain, it has been suggested that APO H may be involved in triglyceride (TG) metabolism. In addition to the previously described quantitative polymorphism, we have recently detected a common qualitative polymorphism at the APO H structural locus. To test the role of APO H genetic variation in determining lipoprotein and lipid levels, we have estimated the allelic effects of APO H variation on TG, VLDL, LDL, HDL, HDL3, and total cholesterol on 356 Nigerian blacks (189 males, 167 females). While no significant effect of phenotype was observed on lipoprotein levels, the effect of interaction between phenotype and gender was significant. Therefore, data on males and females were analyzed separately using analysis of variance after adjusting for age and body mass index. Logarithmic transformation of pertinent variables was done to bring the distribution of the variables closer to normality. A statistically significant effect of phenotype was observed on triglyceride levels in females only (P less than 0.05). Further analysis of this phenotypic effect revealed that it is due to the impact of the APO H*3 allele, which raises triglycerides by 9.92 mg/dl as compared to the common allele, APO H*2. These findings are in accordance with the postulated role of APO H in triglyceride metabolism. On the basis of its sex-specific effect, we propose a hypothesis that may explain the combined influence of the quantitative and qualitative polymorphisms at the APO H locus on triglyceride levels in females.
Cholesterol↗
Genetic studies of human apolipoproteins. X. The effect of the apolipoprotein E polymorphism on quantitative levels of lipoproteins in Nigerian blacks.
Human apolipoprotein E exhibits genetic polymorphism in all populations examined to date. By isoelectric focusing and immunoblotting, three common alleles have been demonstrated in 365 unrelated Nigerian blacks. Furthermore, the APO E genetic polymorphism's effect on quantitative levels of lipids and lipoproteins has been determined. The respective frequencies of the APO E*2, APO E*3, and APO E*4 alleles are .027, .677, and .296. The effect of APO E polymorphism is significant only on total cholesterol and low-density lipoprotein cholesterol. The average excesses of the APO E*2 allele are to lower total cholesterol and low-density lipoprotein cholesterol by 9.19 mg/dl and 11.11 mg/dl, respectively. The average excesses of the APO E*4 allele are to increase total cholesterol and low-density lipoprotein cholesterol by 5.64 mg/dl and 6.18 mg/dl, respectively. On the basis of the differences in (a) the distribution of APO E allele frequencies between the Nigerians and other populations and (b) dietary lipids, we propose a model that shows that lipid metabolism is influenced by the combined effects of the APO E polymorphism and environmental factors.
Alleles↗
Genetic studies of human apolipoproteins. XI. The effect of the apolipoprotein C-II polymorphism on lipoprotein levels in Nigerian blacks.
The human apolipoprotein C-II locus exhibits genetically determined structural polymorphism in United States and African blacks. In the present study, we have investigated the effect of the apoC-II polymorphism on quantitative serum levels of total cholesterol, total high density lipoprotein (HDL) cholesterol, cholesterol in high density lipoprotein subfractions, low density lipoprotein (LDL) cholesterol, and triglycerides (TG) in a sample of 368 unrelated Nigerian blacks. The frequencies of the APOC-II*1 and APOC-II*2 alleles in the samples were 0.947 and 0.053, respectively. In males, the effect of the APOC-II*2 allele was to lower the total serum cholesterol and LDL-cholesterol levels by 13.28 mg/dl and 10.55 mg/dl, respectively, relative to the common allele, APOC-II*1. In females, the effect was to lower total plasma cholesterol by 4.49 mg/dl and LDL-cholesterol by 3.21 mg/dl. The effect of apoC-II on quantitative lipoprotein levels is shown to be independent of variation at the linked apoE locus, but the products of the two loci interact in determining overall quantitative phenotypes.
Adolescent↗
Genetic studies of human apolipoproteins. III. Polymorphism of apolipoprotein C-II.
Using a simple and rapid one-dimensional isoelectric focusing technique followed by immunoblotting, we have detected genetic polymorphism of human apolipoprotein C-II (APO C-II) in normal unfractionated plasma samples of individuals of black ancestry. Two common autosomal codominantly expressed alleles, designated APO C-II*1 and APO C-II*2, at the APO C-II structural locus have been observed with frequencies of 0.975 and 0.025 in US blacks and 0.943 and 0.049 in Nigerian blacks. In addition, the gene product of a rare allele designated APO C-II*3 was observed in a single Nigerian black. Apart from a single example of an APO C-II 2-1 phenotype in plasma samples from 187 whites, which was electrophoretically identical to the 2-1 phenotype observed in blacks, it appears that APO C-II*2 is a unique black marker of potential importance in anthropogenetic and atherosclerosis studies.
Alleles↗
Genetic studies of human apolipoproteins. VII. Population distribution of polymorphisms of apolipoproteins A-I, A-II, A-IV, C-II, E, and H in Nigeria.
Genetic variation at various human apolipoprotein gene loci plays a vital role in modulating lipid metabolism. However, information regarding genetic variation at apolipoprotein loci is scanty, and the available data are largely restricted to Caucasian populations. Using recently developed isoelectric focusing-immunoblotting techniques, we have screened a large number of serum samples from Nigerian blacks to investigate structural variation at six apolipoprotein loci: A-I, A-II, A-IV, C-II, E, and H. With the exception of a single example of a putative APO A-I variant, the APO A-I and A-II loci were found to be monomorphic. Several new variants have been identified at the APO A-IV locus, which are apparently restricted to the black gene pool. By comparison with Caucasians, in whom APO C-II is invariant, four allelic variants have been identified in the Nigerians at the APO C-II structural locus. A common three-allele polymorphism has been observed for APO E, with a striking high frequency of the APO E*4 allele. In addition to three common alleles, the APO H locus is characterized by having an allele marker unique to blacks. The mean heterozygosity at these apolipoprotein loci is higher in Nigerian blacks as compared with the Caucasians.
Alleles↗
Genetic studies of human apolipoproteins. IV. Structural heterogeneity of apolipoprotein H (beta 2-glycoprotein I).
Human beta 2-glycoprotein I has recently been identified as a component of several human plasma lipoprotein fractions and therefore termed as apolipoprotein H. Its metabolic function in lipid metabolism is not known with certainty, though it may be involved in very-low-density-lipoprotein metabolism. Previously, inherited quantitative variation in beta 2-glycoprotein I has been suggested in man. In this investigation, we document the evidence of genetically determined structural polymorphism of apolipoprotein H or beta 2-glycoprotein I by using thin-layer polyacrylamide isoelectric focusing gels followed by immunological identification by double antibody staining. The apolipoprotein H structural locus is characterized by the occurrence of three common alleles in U.S. whites and blacks. The frequency distributions of the three alleles designated APO H1, APO H2, and APO H3 are .059, .882, and .059 in whites and .017, .902, and .068 in blacks, respectively. In addition, the gene product of a fourth allele, APO H4, has been observed at polymorphic frequency in black individuals and may represent a black marker variant. Family data confirm the hypothesis of four alleles at a single APO H gene locus with an autosomal codominant pattern of inheritance.
Alleles↗
The crystal structure of isobutyl 2,3,4-tri-O-acetyl-1-thio-beta-D-xylopyranoside.
Isobutyl 2,3,4-tri-O-acetyl-1-thio-beta-D-xylopyranoside is monoclinic, P21, with a = 10.134(4), b = 7.748(3), c = 11.726(4) A, beta = 96.63(3) degrees, V = 914.55 A3, Z = 2, Dm = 1.262, Dx = 1.264 g . cm-3, mu(MoK alpha) = 226 M-1. The X-ray intensities of 1724 reflections were measured with Nb-filtered MoK alpha radiation (lambda = 0.7107 A) at room temperature. The structure was solved by direct methods, and refined by full-matrix least squares, with anisotropic thermal parameters for the carbon and oxygen atoms and isotropic thermal parameters for the hydrogen atoms, to a final agreement factor of R = 0.08. The molecule has the 4C1(D) conformation, with puckering parameters Q = 0.582 A, theta = 5.6 degrees, phi = 334.7 degrees. The acetyl groups have the planar, (S)-cis configuration most commonly observed. They are oriented, as in many other per-O-acetylated aldopyranosides, with the acetyl planes within +/- 30 degrees of the C-H bond at the ring-carbon atom to which they are attached. Although this is primarily a van der Waals structure, there is some evidence for CH---O hydrogen-bonding.
Carbohydrate Conformation↗
Genetic studies of human apolipoproteins. VI. Common polymorphism of apolipoprotein E in blacks.
Human apolipoprotein E exhibits genetically determined polymorphism with three common alleles, and plays a pivotal role in lipoprotein metabolism. Except in Caucasians, variation at the apolipoprotein E locus has been studied poorly in world populations. Using isoelectric focusing and immunoblotting, we have determined apolipoprotein E phenotypes and gene frequencies in a large number of individuals of black ancestry from the U.S. and Nigeria. The common three allele polymorphism was observed in both populations, with a striking high frequency of the APO E*4 allele as compared to the reported values in other population groups. The frequencies of the three alleles, APO E*2, APO E*3, and APO E*4, respectively, are: 0.034, 0.706, 0.260 in U.S. Blacks and 0.028, 0.662, 0.310 in Nigerian Blacks.
Apolipoproteins E↗