Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0027960 (mole)
 21,279 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Using a synthetic deoxyoligonucleotide mixture constructed for a tryptic peptide of the bovine enzyme as a probe, cDNA coding for the full-length subunit of spermidine synthase was isolated from a human decidual cDNA library constructed on phage lambda gt11. After subcloning into the Eco RI site of pBR322 and propagation, both strands of the insert were sequenced using a shotgun strategy. Starting from the first start codon, which was immediately preceded by a GC-rich region including four overlapping CCGCC consensus sequences, an open reading frame for a 302-amino-acid polypeptide was resolved. This peptide had an Mr of 33,827, started with methionine, and ended with serine. The identity of the isolated cDNA was confirmed by comparison of the deduced amino acid sequence with resolved sequences of the tryptic peptides of bovine spermidine synthase. The coding strand of the cDNA revealed no special regulatory or ribosome-binding signals within 82 nucleotides preceding the start codon and no polyadenylation signal within 247 nucleotides following the stop codon. The coding region, containing a 13-nucleotide repeat close to the 5' end, was longer than, and very different from, that of the bacterial counterpart. This region seems to be of retroviral origin and shows marked homology with sequences found in a variety of human, mammalian, avian, and viral genes and mRNAs. By computer analysis, the first 200 nucleotides of the 5' end of the coding strand appear able to form a very stable secondary structure with a free energy change of -157.6 kcal/mole.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Human spermidine synthase: cloning and primary structure. 234 93

Human bronchial mucin from a patient suffering from chronic bronchitis was solubilized in aqueous solution containing sodium azide and protease inhibitors and purified by Sepharose 4B and 2B column chromatography. The mucin was further purified by cesium bromide density gradient centrifugation. Sodium dodecyl sulfate-polyacrylamide gel (7.5%) electrophoresis of this material showed high-molecular-weight mucin component(s) at the top of the gel. Chemical analysis of this preparation indicated a typical mucin profile of amino acids and carbohydrates. Ion-exchange chromatography resulted in resolution of the purified mucin into neutral and acidic fractions. Comparison of the chemical composition of these two fractions showed higher mole percentage of threonine, serine, sialic acid, and sulfate in the acidic fraction. Chemical deglycosylation of the purified mucin preparation with trifluoromethane sulfonic acid was carried out at 20 degrees C for 3 1/2 h. Sialic acid, fucose, galactose, and N-acetylglucosamine were completely removed, whereas traces of N-acetylgalactosamine were still detected. High-pressure liquid chromatography of the deglycosylated products from native, neutral, and acidic mucin preparations resulted in a principal peptide, P1, with identical amino acid composition. Cyanogen bromide (CNBr) treatment of the peptide P1 from neutral and acidic mucins and subsequent fractionation of the fragments by high-pressure liquid chromatography resulted in similar peptide profiles. The P1 peptide fraction was further subjected to high-pressure liquid chromatography in a second solvent system, which resulted in two peaks, P1a and P1b. Gel filtration of both peptides in 6 M guanidine hydrochloride indicated a single peak with molecular weight of approximately 97 kDa. The amino acid profile of the two peptides was dominated by high levels of threonine, serine, and proline, which combined accounted for nearly 39% of the total residues, and in most respects, the profile resembled that of native mucin. End-group analysis of the peptide P1a indicated a blocked N-terminus, whereas serine was found to be the N-terminal amino acid in the peptide P1b. Rabbit antibodies prepared against the peptide P1 from native tracheal mucin reacted strongly with neutral and acidic mucin as well as the mucin from human colon. Both neutral and acidic human tracheal mucins were immunologically reactive with mouse monoclonal antibody HMPFG-2, which was prepared against human mammary mucin. However, the response of this antibody to human colonic mucin was rather weak.
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PMID:Neutral and acidic human tracheobronchial mucin. Isolation and characterization of core protein. 237 52

We have investigated the interaction of alpha 2-macroglobulin (alpha 2M) with the serine proteinase urokinase, an activator of plasminogen. Urokinase formed sodium dodecyl sulfate stable complexes with purified alpha 2M and with alpha 2M in plasma. These complexes could be visualized after polyacrylamide gel electrophoresis by protein blots using 125I-labeled anti-urokinase antibody or by fibrin autography, a measure of fibrinolytic activity. According to gel electrophoretic analyses under reducing conditions, urokinase cleaved alpha 2M subunits and formed apparently covalent complexes with alpha 2M. Urokinase cleaved only about 60% of the alpha 2M subunits maximally at a mole ratio of 2:1 (urokinase: alpha 2M). Binding of urokinase to alpha 2M protected the urokinase active site from inhibition by antithrombin III-heparin and inhibited, to a significant extent, plasminogen activation by urokinase. Reaction of urokinase with alpha 2M caused an increase in intrinsic protein fluorescence and, thus, induced the conformational change in alpha 2M that is characteristic of its interactions with active proteinases. Our results indicate that both in plasma and in a purified system the alpha 2M-urokinase reaction is functionally significant.
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PMID:Structural and functional characterization of the inhibition of urokinase by alpha 2-macroglobulin. 241 80

Phosphorylation of histone H1 occurs when spermatozoa of the sea urchin Strongylocentrotus purpuratus are treated with the macromolecular fraction of solubilized egg jelly. Phosphorylation is on serine residues in the N-terminal fragment of H1 bisected with N-bromosuccinimide. Phosphorylation is maximal by 4-8 min and dependent on Ca2+, but independent of Na+ or increased intracellular pH. Phosphorylation of H1 can be dissociated from the induction of the acrosome reaction. Only a fraction of the H1 molecules become phosphorylated upon treatment of sperm with egg jelly. The amount of phosphate per mole of H1 increases from 0.15 moles before jelly treatment to 0.46 moles after maximal phosphorylation. Phosphorylation of H1 occurs in a cAMP-dependent manner as indicated by the ability of the phosphodiesterase inhibitors IBMX and SQ20009 to induce H1 phosphorylation. This phosphorylation reaction can be blocked by digesting the sperm surface with Pronase, or preincubation of sperm in wheat germ agglutinin, showing that a ligand in egg jelly must interact with a sperm surface receptor to activate the kinase phosphorylating H1.
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PMID:Phosphorylation of sperm histone H1 is induced by the egg jelly layer in the sea urchin Strongylocentrotus purpuratus. 242 45

Following incubation of HPV 1-induced warts in the presence of [32P] phosphate several of the E4-encoded proteins were found to be radiolabeled. Two-dimensional isoelectric focusing sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicated that the 17K E4 polypeptides had incorporated [32P]phosphate whereas those of 16K were unlabeled. Purified E4 gene products were separated by ion exchange chromatography into a large number of different species, which were of similar size but of different charge due to varying extents of phosphorylated peptides have been isolated and identified. Phosphoserine and phosphothreonine were identified in all 16/17K E4 fractions but not phosphotyrosine. Both HPV 1 E4 16K and 17K fractions were phosphorylated in vitro by cAMP-dependent protein kinase but not by myosin light chain kinase or by phosphorylase kinase. Incubation with cAMP PK gave incorporation of approx. 0.5 mole phosphate/mol of protein indicating that the cAMP-dependent protein kinase site(s) was partially phosphorylated in vivo. This view was supported by the fact that species which were more heavily phosphorylated in vivo incorporated less phosphate after cAMP-dependent protein kinase phosphorylation. HPV 1 E4 was also phosphorylated at serine and threonine residues by a crude cytoplasmic extract prepared from cultured human keratinocytes and cultured human retinoblasts. These results are discussed in the light of the known effects of phosphorylation on the interactions of other keratinocyte-specific proteins.
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PMID:Phosphorylation of the human papillomavirus type 1 E4 proteins in vivo and in vitro. 247 Jan 93

The voltage-sensitive sodium channel from the electroplax of Electrophorus electricus is selectively phosphorylated by the catalytic subunit of cyclic-AMP-dependent protein kinase (protein kinase A) but not by protein kinase C. Under identical limiting conditions, the protein was phosphorylated 20% as rapidly as the synthetic model substrate kemptamide. A maximum of 1.7 +/- 0.6 equiv of phosphate is incorporated per mole. Phosphoamino acid analysis revealed labeled phosphoserine and phosphothreonine at a constant ratio of 3.3:1. Seven distinct phosphopeptides were identified among tryptic fragments prepared from radiolabeled, affinity-purified protein and resolved by HPLC. The three most rapidly labeled fragments were further purified and sequenced. Four phosphorylated amino acids were identified deriving from three consensus phosphorylation sites. These were serine 6, serine 7, and threonine 17 from the amino terminus and a residue within 47 amino acids of the carboxyl terminus, apparently serine 1776. The alpha-subunits of brain sodium channels, like the electroplax protein, are readily phosphorylated by protein kinase A. However, these are also phosphorylated by protein kinase C and exhibit a markedly different pattern of incorporation. Each of three brain alpha-subunits displays an approximately 200 amino acid segment between homologous repeat domains I and II, which is missing from the electroplax and skeletal muscle proteins [Noda et al. (1986) Nature (London) 320, 188; Kayano et al. (1988) FEBS Lett. 228, 1878; Trimmer et al. (1989) Neuron 3, 33]. Most of the phosphorylation of the brain proteins occurs on a cluster of consensus phosphorylation sites located in this segment. This contrasts with the pattern of highly active sites on the amino and carboxyl termini of the electroplax protein. The detection of seven labeled tryptic phosphopeptides compared to the maximal labeling stoichiometry of approximately 2 suggests that many of the acceptor sites on the protein may be blocked by endogenous phosphorylation.
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PMID:Identification of phosphorylation sites for adenosine 3',5'-cyclic phosphate dependent protein kinase on the voltage-sensitive sodium channel from Electrophorus electricus. 255 2

Pigeon and chicken skeletal muscle phosphorylase kinase purified to a nearly homogeneous state is able to phosphorylate both cardiac and skeletal troponin I and T. After 1-hr incubation, the enzyme transfers up to 0.35 mole of phosphorus per mole of skeletal troponin I, up to 0.5 mole of cardiac troponin I and up to 0.1 mole of cardiac and skeletal troponin T. Avian muscle phosphorylase kinase does not phosphorylate the first serine residue of cardiac and skeletal troponin T, but catalyzes the phosphate incorporation into the site(s) of troponin T located in the central or C-terminal parts of the protein molecule. The rate of troponin phosphorylation by pigeon muscle phosphorylase kinase is pH-dependent: the 6.8/8.2 ratio for troponin I is close to 0,2, whereas that with troponin T varies in the range of 0.5-0.7. Troponin phosphorylation by avian phosphorylase kinase depends on the presence of Ca2+ in the incubation mixture. In the presence of 3 mM EGTA troponin I phosphorylation is inhibited by 70-90%, whereas that of troponin T--by 50%. The experimental results indicate that the phosphorylation of troponin I and T is catalyzed either by two different active centers or by different conformations of the single center of avian phosphorylase kinase.
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PMID:[Phosphorylation of isolated components of the troponin complex of skeletal and cardiac muscle phosphorylase kinase from bird skeletal muscles]. 259 Jun 82

The binding of Congo red to several purified amyloid-like peptides having a beta-pleated sheet conformation was quantitatively examined. Congo red binds preferentially to the beta-pleated sheet conformation of both insulin fibrils and poly-L-lysine. Congo red does not bind nearly so well to poly-L-serine or polyglycine, despite the fact that these peptides also have a beta-pleated sheet conformation. Binding to insulin fibrils was saturable with an apparent Bmax of 2 moles of Congo red per mole of insulin fibrils and an apparent KD of 1.75 x 10(-7) M. Binding to beta-poly-L-lysine was similar but had a much higher apparent Bmax of 43. Binding of Congo red to beta-poly-L-lysine was pH dependent and appeared to be determined by the number of protonated lysine residues in the 250 amino acid peptide. We present a new hypothesis in which Congo red binds to amyloid-like proteins via bonds between the two negatively charged sulfonic acid groups of Congo red and two positively charged amino acid residues of two separate protein molecules which are properly oriented by virtue of the beta-pleated sheet conformation of the peptide backbone.
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PMID:Quantitative evaluation of congo red binding to amyloid-like proteins with a beta-pleated sheet conformation. 266 10

Exchangeable phospho- and sphingolipid probes (phosphatidylcholine, -ethanolamine, -serine, and -glycerol, phosphatidic acid, sphingomyelin, cerebroside, and sulfatide) have been synthesized in which one acyl chain is substituted with a fluorescent bimanyl, 7-(dimethylamino)coumarin-3-yl, or diphenyl-hexatrienyl group. The distribution of these probes between two different populations of lipid vesicles can be readily monitored by fluorescence intensity measurements, as described by Nichols and Pagano [Nichols, J. W., & Pagano, R. E. (1982) Biochemistry 21, 1720-1726], when one of the vesicle populations contains a low mole fraction of a nonexchangeable quencher, (12-DABS)-18-PC. The probes examined in this study exchange between phospholipid vesicles on a time scale of minutes, with kinetics indicating that the transfer process takes place by diffusion of probe monomers through the aqueous phase. As expected, lipid probes with different charges differ markedly in their equilibrium distributions between neutral and charged lipid vesicles. However, probes with different polar headgroups differ only modestly in their relative affinities for vesicles composed of "hydrogen-bonding" lipids (PE and PS) vs "non-hydrogen-bonding" lipids (PC and PG or O-methyl-PA). Probes with different headgroups also show modest, albeit reproducible, differences in their relative affinities for cholesterol-containing vs cholesterol-free PC/PG vesicles. Our results suggest that lipids with different headgroup structures may mix more nearly ideally in liquid-crystalline lipid bilayers than would be predicted from previous analyses of the phase diagrams for binary lipid mixtures.
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PMID:Partitioning of exchangeable fluorescent phospholipids and sphingolipids between different lipid bilayer environments. 271 53

Paramyosin from Caenorhabditis elegans was examined for post-translational modification by phosphorylation. Paramyosin purified from populations of mixed-age animals contained 0.7 to 2.0 moles of phosphate per mole of paramyosin. Paramyosin was also phosphorylated in vitro by an endogenous kinase in the particulate fraction. Analysis of the in vitro phosphorylated paramyosin in comparison with the DNA sequence of the unc-15 paramyosin gene of C. elegans shows that serine residues in the non-alpha-helical N-terminal region are the targets of the kinase. The N-terminal region of paramyosin has significant similarity to the non-helical C-terminal region of the two body wall myosin heavy chains of C. elegans. All three regions contain three copies of a Ser-*-Ser-*-Ala motif, the most likely target for phosphorylation in paramyosin, suggesting that these regions may be modified by the same kinase.
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PMID:Phosphorylation of the N-terminal region of Caenorhabditis elegans paramyosin. 275 33


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