Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0019204 (hepatocellular carcinoma)
 71,386 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Hepatic triglyceride lipase (H-TGL) was isolated from human postheparin plasma by column chromatography on heparin-Sepharose and phenyl-Sepharose and immunoaffinity chromatography with monoclonal antibodies. The purified enzyme had an apparent molecular weight of 65,000 on sodium dodecyl sulfate-polyacrylamide gel electrophoresis and an amino-terminal sequence of Leu-Gly-Gln-Ser-Leu-Lys-Pro-Glu. Partial amino acid sequences of seven cyanogen bromide peptides were obtained. A human hepatoma cDNA library was screened with synthetic oligonucleotides derived from the partial protein sequence. The cloned H-TGL cDNA of 1569 nucleotides predicts a mature protein of 477 amino acids plus a leader sequence of 22 amino acids. Blot hybridization analysis of poly(A)+ mRNA with a putative H-TGL cDNA clone gave a single hybridizing band of 1.7 kilobases. The protein contains four consensus N-glycosylation sequences based on the cDNA sequence. Comparison of the enzyme sequence with that of other lipases reveals highly conserved sequences in regions of putative lipid and heparin binding. The carboxyl terminus of H-TGL contains a highly basic sequence which is not reported to be present in rat H-TGL or other members of the lipase gene family.
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PMID:Isolation and cDNA sequence of human postheparin plasma hepatic triglyceride lipase. 283 10

Immunoadsorbents were obtained by coupling antibodies to Sepharose 4 B activated with cyanogen bromide. Thus immobilized antibody directed against bovine kidney gamma-glutamyltransferase was used for immunoaffinity chromatography of the enzyme from bovine kidney and liver, from cow milk and from sheep kidney and liver. Immobilized anti-rat kidney gamma-glutamyltransferase antibody was used for purification of the enzyme from rat kidney, mouse kidney, hamster kidney and rat Morris hepatoma 5123D. Yields of the protease-solubilized gamma-glutamyltransferases isolated on immunoadsorbents columns were usually over 60%. The purified enzymes were almost homogenous in polyacrylamide gel electrophoresis. The enzymes showed different molecular weights and electrophoretic mobilities. The effect of antibodies on affinity of the enzymes to substrate and inhibition by synthetic anthglutin and its isomers were studied.
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PMID:Purification of gamma-glutamyltransferases by immunoaffinity chromatography and some properties of the enzymes. 287 22

A study was made of the ouabain effect (10(-3] on cell proliferation and the dependence of ATP hydrolysis on Na/K-concentration in homogenates of mouse hepatoma (XXIIa) and of L-cells, both sensitive and resistant to etidium bromide. Na+, K+-ATPase activity was found in homogenates of cells from sparse cultures in the presence of ouabain, the activity being stimulated by the Na/K-ratio pecular for the maximum enzymatic activity in cells from the dense cultures. The effect of ouabain on the cell proliferation is similar to the effect of transition of sparse cultures to dense ones.
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PMID:[Effect of ouabain on the Na+, K+-ATPase function and multiplication of transformed cells in culture]. 300 84

The X-chromosome-linked glucose-6-phosphate dehydrogenase (D-glucose-6-phosphate:NADP+ oxidoreductase, EC 1.1.1.49) of humans and other mammals consists of a subunit with a molecular weight of about 58,000. The enzyme plays a key role in the generation of NADPH, particularly in matured erythrocytes, and the genetic deficiency of the enzyme is associated with chronic and drug- or food-induced hemolytic anemia in humans. The enzyme was purified to homogeneity from human erythrocytes. The complete amino acid sequence of the subunit, consisting of 531 amino acid residues, was determined by automated and manual Edman degradation of tryptic, chymotryptic, thermolytic, and cyanogen bromide peptides obtained from the enzyme. Based on the amino acid sequence data thus obtained, a 41-mer oligonucleotide with unique sequence was prepared. Two cDNA libraries constructed in phage lambda gt11--i.e., a human liver cDNA library and a human hepatoma Li-7 cDNA library--were screened with the synthetic nucleotide probe. Two positive clones, lambda G6PD-19 and lambda G6PD-25, were obtained from the hepatoma library. lambda G6PD-19 contained an insertion of 2.0 kilobase pairs (kbp), and encoded 204 amino acid residues that were completely compatible with the COOH-terminal portion of the enzyme. The insertion of the clone had a 3' noncoding region of 1.36 kbp. The other clone, lambda G6PD-25, had an insertion of 1.8 kbp and encoded 362 amino acid residues of G6PD. Southern blot analysis of DNA samples obtained from cells with and without the human X chromosome indicated that the cDNA hybridizes with a sequence in the X chromosome.
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PMID:Human glucose-6-phosphate dehydrogenase: primary structure and cDNA cloning. 301 56

The isolation and characterization of a human apolipoprotein B 100-specific cDNA clone (lambda gt-B1) containing a 1321 base pairs (bp) spanning insert is described. It encodes the 3'-nontranslated 281 bp long region up to the polyadenylation site and 1040 bp of the C-terminal coding region of 345 amino-acid residues of human apo B 100 and the stop codon. The lambda gt-B1 cDNA clone has been isolated from a human hepatoma cDNA expression library by immunoscreening using affinity-purified polyclonal anti apo B 100 antibodies. The nucleotide sequence of the apo B 100 insert has been determined. A part of the polypeptide sequence derived from this nucleotide sequence was identical with the amino-acid sequence obtained by protein sequencing of a purified cyanogen bromide fragment of apo B 100. The fusion protein consisting of beta-galactosidase and the 345 amino-acid residue long C-terminus of apo B 100 had an apparent molecular mass of 148 kDa in NaDodSO4 polyacrylamide gel electrophoresis. In Northern blot hybridization analysis the insert of the apo B 100-cDNA clone hybridized to a 20 to 22 kb mRNA from adult human liver.
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PMID:Isolation, expression and characterization of a human apolipoprotein B 100-specific cDNA clone. 302 65

We describe a rapid and direct assay of NAD(P)H:(quinone-acceptor) oxidoreductase (EC 1.6.99.2) activity in cultured cells suitable for identifying and purifying inducers of this detoxication enzyme. Hepa 1c1c7 murine hepatoma cells are plated in 96-well microtiter plates, grown for 24 h, and exposed to inducing agents for another 24 h. The cells are then lysed and quinone reductase activity is assayed by the addition of a reaction mixture containing an NADPH-generating system, menadione (2-methyl-1,4-naphthoquinone), and MTT [3-(4,-5-dimethylthiazo-2-yl)-2,5-diphenyltetrazolium bromide]. Quinone reductase catalyzes the reduction of menadione to menadiol by NADPH, and MTT is reduced nonenzymatically by menadiol resulting in the formation of a blue color which can be quantitated on a microtiter plate absorbance reader. The reaction is more than 90% dicoumarol inhibitable and menadione dependent. The results are comparable to those obtained by harvesting cells from larger plates, preparing cytosols, and carrying out spectrophotometric measurements.
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PMID:Direct measurement of NAD(P)H:quinone reductase from cells cultured in microtiter wells: a screening assay for anticarcinogenic enzyme inducers. 338 6

By a combination of protein chemistry and recombinant DNA methods a glycine-rich region was found to be located near the carboxyl terminus of the nucleolar specific phosphoprotein, nucleolin, from Novikoff hepatoma (protein C23) and Chinese hamster ovary cells (100-kDa nucleolar protein). A sequence of 192 amino acid residues was derived from partial sequences of cyanogen bromide and N-bromosuccinimide fragments of protein C23 and deduced protein sequence from Chinese hamster ovary cell 100-kDa cDNA sequences. The 66 residues sequenced by protein methods were identical to the corresponding residues deduced by DNA sequencing. The multiple residues of NG,NG-dimethylarginine (DMA) contained in the nucleolin polypeptide were found to be limited to a segment of less than 10 kDa near the carboxyl-terminal end of the protein. This segment also contained internally repeated sequences (e.g. 7 copies of the sequence Gly-Gly-Arg-Gly-Gly were found) which were unrelated to sequences closer to the amino-terminal end. Most arginine residues in this region were surrounded by 2 or 3 glycine residues and were relatively close in sequence to phenylalanine residues.
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PMID:Protein and cDNA sequence of a glycine-rich, dimethylarginine-containing region located near the carboxyl-terminal end of nucleolin (C23 and 100 kDa). 375 37

A large proportion of the metabolites formed from benzo[a]pyrene (BP) in cell cultures from rodents, fish and humans result from conjugation of an oxidized metabolite of BP with sulfate, glucuronic acid or glutathione (GSH). To improve the analysis of these metabolites, a reversed-phase ion-pair h.p.l.c. system using a step gradient of methanol:tetrabutyl-ammonium bromide in ammonium formate buffer has been developed for the separation of these three classes of conjugates. This system separated 3-hydroxy-BP glucuronide and sulfate conjugates and resolved them from GSH conjugates of BP 4,5-oxide, 7,8-oxide and 7,8-diol-9,10-epoxide. Cultures of early passage Syrian hamster, Wistar rat and Sencar mouse embryo cells, a bluegill fry (BF-2) cell line and a human hepatoma cell line (HepG2) were exposed to [3H]BP for 24 h. Medium samples from each were extracted with chloroform: methanol:water, and the water-soluble metabolites were analyzed by ion-pair h.p.l.c. The largest peak of metabolites in the media from cell cultures from rodents and the bluegill fry cell line co-eluted with the glucuronic acid conjugate of 3-hydroxy-BP. These phenol-glucuronides represented 48-62% of the total water-soluble metabolites in the fish and rodent cell cultures. Treatment of this material with beta-glucuronidase released 3-hydroxy-BP and 9-hydroxy-BP in ratios from 3:4 to 13.3:1 in various cultures. Media from the bluegill fry cell line and the mouse embryo cell cultures also contained a peak of BP-diol glucuronides; treatment of these peaks with beta-glucuronidase released mainly BP-7,8-diol. In HepG2 cells, 40% of the water-soluble metabolites were identified as sulfate conjugates of 3-hydroxy-BP and 9-hydroxy-BP. No glucuronic acid conjugates of BP metabolites were detected in HepG2 cells. Only small amounts of the water-soluble metabolites from these cell cultures eluted in the same volumes as the synthetic GSH conjugate of BP-4,5-oxide, BP-7,8-oxide and BP-7,8-diol-9,10-oxide. These studies indicate that conjugation with glucuronic acid represents a major pathway of formation of water-soluble metabolites from BP in cells derived from a number of species and demonstrate the value of this ion-pair h.p.l.c. system for the analysis of conjugates formed from BP.
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PMID:Separation by ion-pair high-performance liquid chromatography of the glucuronide, sulfate and glutathione conjugates formed from benzo[a]pyrene in cell cultures from rodents, fish and humans. 380 96

Regulation of synthesis of carbamoyl-phosphate synthetase I by glucocorticoids, 8-bromoadenosine 3',5'-monophosphate (8-bromo-cAMP), and insulin was investigated in Reuber hepatoma H-35. By measuring the incorporation of [35S]methionine into carbamoyl-phosphate synthetase I and its precursor, we showed that dexamethasone stimulates the enzyme synthesis approximately fivefold. A detectable stimulation was observed at 1 nM of dexamethasone, half-maximal stimulation at 4 nM, and maximal stimulation above 40 nM. Corticosterone was more effective than dexamethasone both for the minimal concentration needed and for the extent of the stimulation. Hydrocortisone was less effective than dexamethasone. 8-Bromo-cAMP also stimulated the enzyme synthesis at a concentration of 3 mM. The effect of 8-bromo-cAMP was suggested to be additive to the effect of dexamethasone. Physiological concentrations of insulin strongly suppressed the stimulatory effect of dexamethasone on the enzyme synthesis but could not completely counteract the effect of dexamethasone. The half-maximal and maximal effects of insulin were observed at 0.5 nM and 5 nM, respectively. Insulin also counteracted the effect of 8-bromo-cAMP on the enzyme synthesis.
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PMID:Interaction between glucocorticoids, 8-bromoadenosine 3',5'-monophosphate, and insulin in regulation of synthesis of carbamoyl-phosphate synthetase I in Reuber hepatoma H-35. 389 22

Regiospecific syntheses of gamma- and alpha-conjugates of methotrexate and poly(L-lysine) are described. The alpha- and gamma-t-butyl esters, respectively, of methotrexate were coupled to poly(L-lysine) with diphenylphosphoryl azide in N,N-dimethylformamide, the ester-protecting group was cleaved with 15% hydrogen bromide in acetic acid, and small molecules were removed by dialysis. Poly(L-lysine) of Mr = 1,500-8,000 and 8,000-30,000 was used to prepare six different conjugates, which were characterized by ultraviolet absorbance measurement and quantitative amino acid analysis. The degree of substitution varied from one methotrexate per 4.7 lysines to one methotrexate per 10.2 lysines. Dihydrofolate reductase inhibition in a cell-free assay was observed with alpha- and gamma-conjugates, but the latter had the greater affinity (only 3-fold less than that of methotrexate itself). The binding of the conjugates exhibited a slight pH dependence, with affinity being greater at pH 7.2 than at pH 8.5 for both alpha- and gamma-conjugates. Toxicity to cultured rat hepatoma cells (H35) was also greater for the gamma-conjugates, and showed some dependence on the chain-length and degree of substitution of the poly(L-lysine) carrier. Cells resistant to methotrexate by virtue of a transport defect (H35R0.3 line) retained their sensitivity to the gamma-conjugate, but less so to the alpha-conjugate. There was also some retention of sensitivity in a more highly resistant cell line (H35R10) with impaired methotrexate transport and a concomitant increase in dihydrofolate reductase activity. gamma-Conjugation was likewise more favorable in cytotoxicity assays against L1210 murine leukemia cells, and there was partial retention of activity against highly methotrexate-resistant lines (L1210/R71 and L1210/R81) with a transport defect and/or an elevation of dihydrofolate reductase content. In antitumor assays against intraperitoneal L1210 leukemia in mice, a gamma-conjugate with Mr = 8,000-30,000 and one methotrexate per 5.5 lysines produced a 35-75% increase in lifespan when administered intraperitoneally at single doses equivalent to 10-20 mg/kg of methotrexate. A similar increase in lifespan with methotrexate alone on the single-dose regimen required 50-150 mg/kg. An alpha-conjugate of similar Mr and degree of substitution was inactive at nontoxic doses, as were other gamma-conjugates of lower Mr and/or degree of substitution.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Regiospecific gamma-conjugation of methotrexate to poly(L-lysine). Chemical and biological studies. 396 26


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