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11.
Isolation and characterization of a Mr = 110,000 glycoprotein localized to the hepatocyte bile canaliculus 总被引:2,自引:0,他引:2
J K Petell M Diamond W J Hong Y Bujanover S Amarri K Pittschieler D Doyle 《The Journal of biological chemistry》1987,262(30):14753-14759
A Mr = 110,000 glycoprotein, GP 110, was partially purified using wheat germ agglutinin-Sepharose affinity chromatography from a bile canalicular-enriched membrane fraction denoted N2u of rat liver. This fraction was subjected to preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis and the Mr = 110,000 polypeptide was excised and used as an immunogen in rabbits. The antisera were found to specifically recognize a Mr = 110,000 polypeptide, named GP 110, in the N2u membrane fraction. In isolated hepatocytes, GP 110 was readily accessible to cell surface iodination catalyzed by lactoperoxidase at 4 degrees C and was judged by immunoprecipitation studies to contain about 2% of total radioactivity incorporated into externally oriented proteins of the cell. Immunoprecipitated GP 110 was shown by two-dimensional polyacrylamide gel electrophoresis to migrate with an approximate pI of 4.9. Indirect immunofluorescence on frozen liver sections demonstrated that GP 110 was primarily localized in the bile canaliculus. In corroborative studies employing subcellular fractionation, it was found that GP 110 was enriched nearly 19-fold in P2, a plasma membrane fraction primarily derived from the sinusoidal domain, and 44-fold in N2u. In contrast, only low levels of GP 110 were present in endoplasmic reticulum, mitochondrial, cytosolic, and nuclear-enriched fractions of liver. The physiological function of GP 110 is as yet unknown; antisera to it did not immunoprecipitate other known bile canalicular proteins of similar molecular weights. GP 110 was found to be extensively glycosylated relative to other known membrane proteins; approximately 33% of the apparent molecular weight appear to be carbohydrate. In agreement, limited removal of N-linked carbohydrate chains indicated that there are approximately eight chains/GP 110 polypeptide. Neuraminidase treatment of GP 110 resulted in a desialylated Mr = 85,000 polypeptide suggesting that the majority of carbohydrate chains on GP 110 are of the complex type. 相似文献
12.
T C Madhok C C Chao S G Matta A Hong B M Sharp 《Biochemical and biophysical research communications》1989,165(1):151-157
Polyclonal antibodies were raised against a synthetic decapeptide (designated S3) predicted from a segment of the alpha-3 subunit cDNA (amino acid residues 130-139) encoding the rat brain nicotinic cholinergic receptor. This segment was selected because it may be proximate to the nicotine/acetylcholine-binding site of the receptor (1). By radioligand binding assays and sucrose density gradient centrifugation, these monospecific antibodies were shown to inhibit the binding of [3H]nicotine to both the large molecular weight rat brain receptor (240 kDa) and to an SDS-disaggregated nicotine-binding subunit species (80 kDa), in a dose-dependent manner. The neutralizing effect of the anti-S3 antibodies supports the view that this region of the protein is closely related to the agonist binding site. 相似文献
13.
人体小卫星DNA探针的制备 总被引:3,自引:2,他引:1
根据人体小卫星DNA核心顺序,化学合成长23碱基寡核苷酸探针,筛选人体基因组文库,旨在获得能用作遗传分析探针的小卫星顺序。结果得到15个含小卫星的阳性重组子。随机取其一(C_(35.9))作探针,试做群体分析。所有个体均可检出多条杂交带。其中某些带具有多态性。在一定检测条件下,检出的DNA图谱在有限的个体内具有个体特异性。结果表明筛选文库得到的小卫星顺序可用于小卫星多态性的检测。其它小卫星探针的筛选和应用性研究正在进行。 相似文献
14.
15.
Optimal substrate feeding policy for a fed batch fermentation with substrate and product inhibition kinetics 总被引:1,自引:0,他引:1
Hong J 《Biotechnology and bioengineering》1986,28(9):1421-1431
The optimal substrate feeding policy for the fed batch fermentation which is governed by product and substrate inhibited kinetics is presented. The conjunction point between nonsingular and singular arcs and the feeding policy along the singular arc are derived analytically in terms of the concentrations of substrate and product and the liquid volume. Thus, it is possible to determine the feeding rate by monitoring the state variables (i.e., closed loop control). As a specific example, an optimization study of the fed batch fermentation for ethanol production by Saccharomyces cerevisiae is presented. It is shown that the optimal feeding patterns are heavily dependent upon the initial conditions. The point selectivity provides the guideline for predicting the optimal feeding patterns and explaining the results of rigorous mathematical analysis. 相似文献
16.
Isolation of a cDNA for human muscle 6-phosphofructokinase 总被引:1,自引:0,他引:1
A cDNA for human muscle 6-phosphofructokinase (EC.2.7.1.11) has been isolated from a human fibroblast cDNA library made using the Okayama-Berg procedure. The cDNA isolated as a Bam H1 fragment of the pcD recombinant, pO4, is approximately 2000 bp in length. It represents approximately 1350 bp of the C-terminus coding sequence of the enzyme, approximately 500 bp of the 3'-untranslated region and approximately 150 bp of the vector sequences. The identity of the pO4 cDNA was established by the observation of a high degree of homology (approximately 95%) between the deduced amino acid sequence with the published protein sequence of rabbit muscle 6-phosphofructokinase, and the assignment of the sequence to human chromosome 1 (the known location of PFKM) by using somatic cell hybrids. Based on immunochemical evidence, we had previously predicted not only a remarkable structural conservation of the vertebrate muscle PFK, but also partial structural identity among all three vertebrate PFK isozymes. The pO4 cDNA is, therefore, expected to permit isolation of cDNAs for muscle and non-muscle PFKs from a wide variety of vertebrate species. 相似文献
17.
Athymic (nude) mice were transplanted with cultured thymic fragments from syngeneic, allogeneic, and partially allogeneic (recombinant) mice. Lymphocyte proliferation and cytotoxicity in vitro were measured to assess immunologic reconstitution. Transplanted nude mice were immunocompetent whether donor and recipient were disparate for class I, class II, or both H-2 gene types. Furthermore, allotolerance for thymic H-2 class I antigens was achieved independently of class II antigen allotolerance. Class I antigen tolerance was not broken during lymphocyte responses to unrelated alloantigens, ruling out insufficient help as the tolerance mechanism. Splenocytes, isolated from nude mice transplanted with fully allogeneic or syngeneic thymic fragments and stimulated in vitro with trinitrophenyl-modified cells, displayed H-2-restricted, hapten-specific cytotoxicity. Cytotoxic cells from allotolerant mice were restricted to either host or thymic H-2 antigens, depending on the stimulating cell haplotype. Response levels for thymic and host trinitrophenyl-modified cells were comparable. We have shown that allogeneic thymic epithelium transplanted into adult nude mice can induce allotolerance to class I and II H-2 antigens equally, and permits T lymphocyte interaction with cells bearing thymic donor or host H-2 antigens. Our results are consistent with a model wherein T lymphocyte self-receptors retain their genomic repertoire but can be selectively mutated or expanded by appropriate H-2 antigen presentation by the thymus. 相似文献
18.
Male, Fischer strain 344 adult rats were given various doses (25-100 mg/kg) of p,p'-DDT by oral gavage, and levels of biogenic amines, their metabolites, and amino acid neurotransmitters, tremor activity, and rectal temperature were measured at several intervals (2, 5, 12, and 24 h) after dosing. Dose-related increases in rectal temperature and in tremor activity were observed at 50-100 mg/kg 12 h after dosing. Tremorigenic doses of DDT increased the 5-hydroxyindoleacetic acid (5-HIAA) level in hypothalamus, brainstem, and striatum, whereas doses of 75 and 100 mg/kg increased the 3-methoxy-4-hydroxyphenylglycol (MHPG) level in hypothalamus and brainstem and the 3,4-dihydroxyphenylacetic acid level in striatum. Six amino acids were assayed in the brainstem, hypothalamus, and striatum; aspartate and glutamate levels were increased only in brainstem at 25-100 mg/kg. No consistent changes in concentrations of taurine, glutamine, glycine, or gamma-aminobutyric acid were observed in any of the regions assayed. Time-related increases in rectal temperature were seen 2-12 h after dosing, and the presence of tremor was observed 5-12 h after dosing; for both the time of peak effect was at 12 h. The DDT-induced hyperthermia and tremor were associated with dose- and time-related increases in levels of 5-HIAA, MHPG, aspartate, and glutamate. It is suggested that an increase in the turnover rate of 5-hydroxytryptamine (5-HT) may be responsible for the DDT-induced hyperthermia, whereas increases in the metabolism of 5-HT and norepinephrine may be involved in the tremor. 相似文献
19.
20.
A S Abramovitz J Y Hong V Randolph 《Biochemical and biophysical research communications》1983,117(1):22-29
We have previously reported the purification of polypeptides from soybean which are potent inhibitors of superoxide production by human neutrophils. We now report that neither oxygen uptake nor hydrogen peroxide production by stimulated neutrophils is affected by these inhibitors. Furthermore, the E-1 and E-3 polypeptides inhibit ferricytochrome c reduction by a xanthine oxidase superoxide generation system. The inhibitory activity of E-3 in the model system is blocked by 1 mM KCN while E-1 is only slightly cyanide sensitive. Atomic absorption analysis of E-1 and E-3 polypeptides reveal copper in the latter and manganese in the former. Thus, E-3 is a copper-containing superoxide dismutase while E-1 appears to be a manganese-containing superoxide dismutase. 相似文献