Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The method of hydroxylapatite-mediated rapid and effective transfer of DNA onto nitrocellulose filters for following dot-hybridization was elaborated. The analysed DNA occurred initially in diluted and large volume solutions (from 1 to 10 ml) with various composition (2 M NaCl; 4 M LiCl--8 M urea; 4 M CsCl; 5 and 20% sucrose) was adsorbed on hydroxylapatite and quantitatively transferred onto nitrocellulose after hydroxylapatite solubilization in a small volume of acid (usually, 200 microliters of 10% TCA). As exemplified by the hybridization of total rat liver DNA with the plasmid ph22 DNA containing a cluster of sea urchin histone genes, the method presented appears to be not only simple and useful for handling multiple probes of diluted DNA solutions with high concentrations of salts, sucrose and urea but also more sensitive than some convenient DNA dot-hybridization methods.
Mol Biol (Mosk)
PMID:[Hydroxyapatite-mediated transfer of DNA on nitrocellulose filters in dot-hybridization experiments]. 282 77

Effects of glutamate on myocardial mechanical function and energy metabolism during 120 min of hypoxia and subsequent reoxygenation were studied in the isolated arterially perfused newborn and adult rabbit hearts. The muscle was perfused with a Krebs-Henseleit (KH) solution or KH solution which contained 1 mM glutamate. Glutamate attenuated the effects of hypoxia on mechanical function and tissue ATP concentration, and enhanced the recovery of mechanical function and tissue ATP during reoxygenation. During hypoxia, glutamate increased tissue succinate and GTP with no change in total lactate and pyruvate production. Trace studies using 14C-glutamate and the tissue homogenate showed that hypoxia increased tissue succinate and inhibited TCA cycle. Additional glutamate produced more CO2 and TCA intermediates in both oxygenated and hypoxic mediums. These data indicate that glutamate increased the rate of ATP production in the hypoxic and reoxygenated heart. This study shows that the improvement of mechanical function and ATP formation in the hypoxic myocardium by glutamate was due to an increase in both oxidative phosphorylation and substrate level phosphorylation. The effect of glutamate on the ATP and GTP production in the newborn heart was not different from the adult.
J Mol Cell Cardiol 1986 Sep
PMID:The effect of glutamate on hypoxic newborn rabbit heart. 287 83

The properties of the antigen recognized by monoclonal antibody FH6 have been analyzed. FH6 was originally generated against a glycolipid, i.e. a difucoganglioside isolated from human colonic adenocarcinoma, and specifically reacts with sialyl Lex-i determinant. Several culture supernatants of human carcinoma cell line cells were found to have high levels of FH6-reactive antigen, and PC-9, a human lung carcinoma cell line was used for the analysis. A solid-phase sandwich radioimmunoassay was performed to detect the antigen. The antigenic activity was extractable in 0.6 M PCA or 7% TCA, and was sensitive to mild alkaline treatment and to Pronase digestion. Most of the antigen was eluted in the void volume of a Sepharose CL-2B column, which indicates that its molecular weight is greater than several million. It was eluted from a DEAE-cellulose column at a NaCl concentration in the range of 0.2-0.25 M. The immunoaffinity-purified antigen has a high carbohydrate content of more than 80%. These data indicate that the antigen recognized by FH6 in the culture supernatant of PC-9 is not a glycolipid, but a high molecular weight glycoprotein which could be referred to as a mucin, or a proteoglycan, which contains keratan-sulfate like glycosaminoglycan chains, as judged from the results of the glycosidase treatments.
J Mol Recognit 1988 Jun
PMID:Glycolipid-directed FH6 monoclonal antibody recognizes high molecular weight glycoprotein antigen carrying sialyl Lex-i determinant in the culture supernatant of PC-9 cells. 290 4

Four Neurospora crassa genomic clones have been selected as hybridizing much more strongly to labelled mRNA isolated from acetate-grown mycelium than to mRNA from sucrose-grown mycelium. Hybridization of restriction fragments with acetate-specific mRNA or cDNA has been used to delimit the transcribed region(s) of each clone. The transcription of all four clones is strongly induced by transfer of growing mycelium from sucrose to acetate as sole carbon source. In wild-type mycelium, mRNAs corresponding to the four clones reach maximum levels after four hours of induction. They accumulate more rapidly and reach higher levels in an acetate non-utilizing mutant, acu-7, which has been found to overproduce enzymes of the glyoxylate cycle and to have a partial block in the TCA cycle. Molecular transformation of a Neurospora acu-5 mutant and of an Aspergillus nidulans acuE mutant by DNA of clone 2 and clone 1, respectively, strongly suggests that clone 2 codes for acetyl-coenzyme A synthetase and that clone 1 codes for malate synthase. The transcribed segments of clones 1 and 2 each hybridize to corresponding clones from Aspergillus nidulans (R. A. Sandeman and M. J. Hynes, personal communication).
Mol Microbiol 1988 Sep
PMID:Molecular cloning, identification and transcriptional analysis of genes involved in acetate utilization in Neurospora crassa. 305 23

An Escherichia coli strain transfected with a plasmid containing four linked human proinsulin genes was grown in the presence of 35S and 3H labelled amino acids to gain access to human insulin that was radiolabelled at 19 evenly distributed sites throughout the amino acid sequence. The multi-proinsulin precursor was cleaved at methionine residues with cyanogen bromide, then the individual proinsulin units were folded via their S-cysteine sulfonate derivative and converted to insulin by enzymatic digestion. Purification steps were carried out by ion-exchange and reverse-phase HPLC techniques. The final radiolabelled biosynthetic human insulin was produced at a specific activity of up to 300 Ci/mmol, and was shown to be indistinguishable from commercially available human insulin according to HPLC behavior, amino acid analysis, immunoreactivity and biological activity. A comparison of the kinetics of processing of 35S/3H-labelled biosynthetic human insulin and 125I-labelled commercial human insulin by murine TA3 hybridoma antigen presenting cells demonstrated that radiolabelled biosynthetic insulin was processed approximately 16 times slower than its iodinated counterpart. Measurable 125I TCA soluble radioactivity was detected extracellularly within 15 min whereas the same amount of extracellular TCA soluble 3H/35S radioactivity was not seen until 240 min. These results begin to address the importance of using a biosynthetically labelled protein as opposed to an iodinated protein to study how an APC handles antigen in a physiological manner.
Mol Immunol 1988 Dec
PMID:Purification and characterization of radiolabelled biosynthetic human insulin from Escherichia coli. Kinetics of processing by antigen presenting cells. 307 Mar 57

125I-labeled ovine follitropin (125I-oFSH) and deglycosylated follitropin (125I-DG-oFSH) were injected into rats and the tissue uptake was quantified and correlated with radioautographic data. Trichloroacetic acid (TCA)-precipitable radioactivity and gel filtration analysis of blood samples indicated no degradation of follitropin or analogue with time. Clearance of follitropin from the circulation was accelerated after its deglycosylation. Disappearance of both molecules from the blood was associated with uptake and/or loss of radioactivity from liver, kidney, ovary and spleen. The more rapid removal of deglycosylated follitropin from blood was associated with higher renal levels of accumulated radioactivity than native follitropin. This was associated with its localization within the cortex, specifically the proximal convoluted tubules of the nephron. Binding of 125I-labeled follitropin and analogue to granulosa cells was specific and time-dependent. 125-I-DG-oFSH demonstrated greater avidity of binding to rat granulosa cells with time than 125I-oFSH. This was associated with slower dissociation kinetics and/or metabolism for 125I-DG-oFSH. The absence of localization of either 125I-follitropin or analogue in hepatic tissue suggests that hepatic mechanisms may not significantly contribute to the clearance of these molecules. Implications of these findings in regard to the metabolism of oFSH and its antagonist are discussed.
Mol Cell Endocrinol 1987 Aug
PMID:Distribution of follitropin and deglycosylated follitropin in the rat: a quantitative and radioautographic study. 311 48

The present study evaluates protein synthesis in rat hippocampal slices maintained in vitro. Transverse slices of hippocampus were prepared from both adult rats and rat pups during postnatal development and incubated in a gassed (95% O2/5% CO2) balanced salt medium containing 5 nM 3H-leucine. The time course of 3H-leucine incorporation into TCA-precipitable protein was determined using slices removed from the media after 5, 10, 20, 30, 40, 60, and 120 min of incubation. The pattern of 3H-amino acid incorporation was evaluated by fixing slices with paraformaldehyde, embedding the slices in plastic, and sectioning the slices end on and en face for autoradiographic analysis. Biochemical analysis of 300 and 400 micron slices revealed that incorporation of leucine into protein proceeds at a constant rate. The autoradiographic analysis revealed that in adult hippocampal slices of 300-600 micron thickness there was complete penetration of 3H-leucine with no indication of a gradient in the extent of incorporation throughout the slice. The pattern of grain density within 300-600 micron slices matches that previously reported after in vivo injections of radiolabeled amino acid, where grain density is highest over neuronal cell bodies and lower over the laminae that contain dendritic processes and axons (Phillips et al: Mol Brain Res 2:251-261, 1987). Hippocampal slices of 200, 800, and 1,000 micron thickness showed irregular labeling. Slices of 200 micron were filled with pyknotic nuclei and vacuoles and exhibited patchy labeling. In 800 micron slices there were isolated areas of good preservation within the slice core, but these areas exhibited little incorporation. Relative to the 300-600 micron slices, there was a higher number of pyknotic nuclei and a much deeper layer of necrosis along the cut edges. Slices of 1,000 micron thickness showed poor preservation throughout and low levels of incorporation. Biochemical studies revealed a much higher rate of incorporation in the slices prepared from postnatal animals. Autoradiography of the slices from developing rats revealed that penetration was excellent and incorporation appeared to be greater as judged by an overall higher grain density. We believe that rat hippocampal slices provide a good in vitro model of protein metabolism that will be useful for studies of protein synthesis in isolated cell body and dendritic laminae and for the evaluation of whether protein synthesis in particular laminae is regulated by synaptic activity.
 ...
PMID:Protein synthesis by rat hippocampal slices maintained in vitro. 321 13

The reproducible photolabeling of the androgen receptor from human skin fibroblasts, using [3H]methyltrienolone (R-1881) as ligand is described. Crude nuclei were irradiated for 2 min using a UV lamp with an emission line at 352 nm and a CuSO4 filter. After KCl extraction, proteins were precipitated with trichloroacetic acid, washed with ether and assayed for radioactivity. Specific binding was determined as the difference in bound radioactivity between cells incubated with [3H]R-1881 +/- a 200-fold excess of unlabeled dihydrotestosterone (DHT). The photolabeled proteins were analyzed on SDS-polyacrylamide gel electrophoresis yielding one peak of 90 kDa and in several cases, one of 43 kDa. These peaks comprised 60 +/- 20% of the saturable binding recovered on the gels. The overall efficiency of photolabeling was between 1 and 5%. The amount of covalently bound radioactivity was proportional to the number of cells used. The labeling was inhibited by R-1881, DHT, the anti-androgens hydroxyflutamide and cyproterone acetate and to a lesser extent by estradiol and progesterone. No covalent attachment of R-1881 to any protein was observed when nuclei from patients with androgen insensitivity were irradiated, whether or not the cells were receptor positive or negative. In conclusion the androgen receptor from human skin fibroblast can be efficiently photolabeled and could be used as a marker to follow receptor purification. The absence of photolabeling of nuclear extracts from receptor-positive androgen-insensitive patients may reflect some abnormality of the receptor.
Mol Cell Endocrinol 1987 Dec
PMID:Photoaffinity labeling of the androgen receptor from human skin fibroblasts. 350 Aug 83

The susceptibility of porcine relaxin and 125I-polytyrosyl-porcine relaxin to degradation by 3 purified enzymes involved in the degradation of insulin and proinsulin was examined. Rat liver glutathione-insulin transhydrogenase (GIT), which cleaves disulfide bonds in insulin, catalyzed a time- and concentration-dependent increase in trichloroacetic acid (TCA)-soluble radioactivity of relaxin. The Sephadex G-50 profile of the reaction products revealed conversion to the A- and B-chains. Relaxin competitively inhibited the degradation of insulin by GIT; however, kinetic analysis revealed insulin to be preferred over relaxin as a substrate. Rat liver cytosol neutral thiol peptidase (NTP) catalyzed a time- and concentration-dependent increase in the TCA solubility of relaxin and a shift in the Sephadex G-50 radioactivity profile to low molecular weight products. Kinetic analysis revealed that insulin and B-chain are preferred over relaxin as substrates for NTP. A third enzyme, rat kidney neutral metalloendopeptidase, which degrades proinsulin and insulin C-peptide but not insulin, also did not degrade porcine relaxin.
Mol Cell Endocrinol 1986 Jun
PMID:Degradation of porcine relaxin by glutathione-insulin transhydrogenase and a neutral peptidase. 351 16

IgM is present in cows milk, is able to bind secretory component (SC), has a purported role as a secretory immunoglobulin in other species and has been identified with various antibody functions in cows milk. To determine the origin of cows milk IgM, we administered extrinsic 131I-IgM to lactating cows with cannulated bile and parotid ducts and studied the kinetics of its disappearance from serum and its appearance in milk, bile and parotid saliva for 60 hr post-injection. Pentameric IgM appeared to require a long equilibration time and disappeared from serum with a T1/2 of 40 hr. The transport of IgM into bile also appeared biphasic. Results showed that no 131I-IgM was transported intact into parotid saliva and that most radioactivity in milk and bile after 6 hr was in the form of low mol. wt, TCA-precipitable fragments rather than of the size of a pentamer. During the first 24 hr only 0.83% of the administered dose reached the milk in pentameric form, nevertheless, isotope dilution calculations indicated that nearly all milk IgM was derived from serum. During a 12 hr collection period, corresponding to one milking, greater than 200 mg of serum IgM is secreted in milk. During the first 24 hr, only 0.70% of the administered IgM reached the bile as a pentamer. It was calculated that 50% of the pentameric IgM in bile, 3 hr after administration, was serum-derived. Twenty-five per cent of the IgM appearing in bile and ca 10% of the IgM appearing milk, becomes associated with secretory component. A hypothesis to explain the degradation associated with this inefficient transport mechanism is presented.
Mol Immunol 1987 Mar
PMID:The transport and metabolism of bovine IgM. 361 12


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