Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: UNIPROT:Q07644 (polypeptide)
72,197 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Lysyl hydroxylase (EC 1.14.11.4), an alpha 2 dimer, catalyzes the formation of hydroxylysine in collagens by the hydroxylation of lysine residues in X-Lys-Gly sequences. We report here on the isolation of cDNA clones coding for the enzyme from a chick embryo lambda gt11 library. Several overlapping clones covering all the coding sequences of the 4-kilobase mRNA and virtually all the noncoding sequences were characterized. These clones encode a polypeptide of 710 amino acid residues and a signal peptide of 20 amino acids. The polypeptide has four potential attachment sites for asparagine-linked oligosaccharides and 9 cysteine residues, at least one of which is likely to be involved in the binding of the Fe2+ atom to a catalytic site. A surprising finding was that no significant homology was found between the primary structures of lysyl hydroxylase and prolyl 4-hydroxylase in spite of the marked similarities in kinetic properties between these two enzymes. A computer-assisted comparison indicated only an 18% identity between lysyl hydroxylase and the alpha-subunit of prolyl 4-hydroxylase and a 19% identity between lysyl hydroxylase and the beta-subunit of prolyl 4-hydroxylase. Visual inspection of the most homologous areas nevertheless indicated the presence of several regions of 20-40 amino acids in which the identity between lysyl hydroxylase and one of the prolyl 4-hydroxylase subunits exceeded 30% or similarity exceeded 40%. Southern blot analyses of chick genomic DNA indicated the presence of only one gene coding for lysyl hydroxylase.
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PMID:Molecular cloning of chick lysyl hydroxylase. Little homology in primary structure to the two types of subunit of prolyl 4-hydroxylase. 170 64

Prolyl 4-hydroxylase (EC 1.14.11.2) catalyzes the formation of 4-hydroxyproline in collagens by the hydroxylation of proline residues in X-Pro-Gly sequences. The reaction requires Fe2+, 2-oxoglutarate, O2, and ascorbate and involves an oxidative decarboxylation of 2-oxoglutarate. Ascorbate is not consumed during most catalytic cycles, but the enzyme also catalyzes decarboxylation of 2-oxoglutarate without subsequent hydroxylation, and ascorbate is required as a specific alternative oxygen acceptor in such uncoupled reaction cycles. A number of compounds inhibit prolyl 4-hydroxylase competitively with respect to some of its cosubstrates or the peptide substrate, and recently many suicide inactivators have also been described. Such inhibitors and inactivators are of considerable interest, because the prolyl 4-hydroxylase reaction would seem a particularly suitable target for chemical regulation of the excessive collagen formation found in patients with various fibrotic diseases. The active prolyl 4-hydroxylase is an alpha 2 beta 2 tetramer, consisting of two different types of inactive monomer and probably containing two catalytic sites per tetramer. The large catalytic site may be cooperatively built up of both the alpha and beta subunits, but the alpha subunit appears to contribute the major part. The beta subunit has been found to be identical to the enzyme protein disulfide isomerase and a major cellular thyroid hormone-binding protein and shows partial homology with a phosphoinositide-specific phospholipase C, thioredoxins, and the estrogen-binding domain of the estrogen receptor. The COOH-terminus of this beta subunit has the amino acid sequence Lys-Asp-Glu-Leu, which was recently suggested to be necessary for the retention of a polypeptide within the lumen of the endoplasmic reticulum. The alpha subunit does not have this COOH-terminal sequence, and thus one function of the beta subunit in the prolyl 4-hydroxylase tetramer appears to be to retain the enzyme within this cell organelle.
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PMID:Protein hydroxylation: prolyl 4-hydroxylase, an enzyme with four cosubstrates and a multifunctional subunit. 253 73

Prolyl 4-hydroxylase [procollagen-proline, 2-oxyglutarate 4-dioxygenase; procollagen-L-proline, 2-oxoglutarate:oxygen oxidoreductase (4-hydroxylating), EC 1.14.11.2], an alpha 2 beta 2 tetramer, catalyzes the formation of 4-hydroxyproline in collagens by the hydroxylation of proline residues in peptide linkages. We report here on the isolation of cDNA clones encoding the alpha-subunit of the enzyme from human tumor HT-1080, placenta, and fibroblast cDNA libraries. Eight overlapping clones covering almost all of the corresponding 3000-nucleotide mRNA, including all the coding sequences, were characterized. These clones encode a polypeptide of 517 amino acid residues and a signal peptide of 17 amino acids. Previous characterization of cDNA clones for the beta-subunit of prolyl 4-hydroxylase has indicated that its C terminus has the amino acid sequence Lys-Asp-Glu-Leu, which, it has been suggested, is necessary for the retention of a polypeptide within the lumen of the endoplasmic reticulum. The alpha-subunit does not have this C-terminal sequence, and thus one function of the beta-subunit in the prolyl 4-hydroxylase tetramer appears to be to retain the enzyme within this cell organelle. Interestingly, three of the cDNA clones for the alpha-subunit contained a 64-nucleotide sequence homologous but not identical to the corresponding 64-nucleotide sequence found in four other cDNA clones. Nuclease S1 mapping experiments demonstrated that this difference was due to the existence of two types of mRNA present in approximately equal amounts. Southern blot analyses of human genomic DNA with a cDNA probe for the alpha-subunit suggested the presence of only one gene encoding the two types of mRNA, which appear to result from mutually exclusive alternative splicing of primary transcripts of one gene.
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PMID:Molecular cloning of the alpha-subunit of human prolyl 4-hydroxylase: the complete cDNA-derived amino acid sequence and evidence for alternative splicing of RNA transcripts. 254 75

Prolyl 4-hydroxylase (EC 1.14.11.2) is a key enzyme required for the posttranslational hydroxylation of proline residues in collagen. The enzyme is a tetramer composed of two pairs of nonidentical subunits (alpha 2 beta 2). The beta subunit is protein disulfide-isomerase, a ubiquitous enzyme found in the endoplasmic reticulum of many cell types. We report here the amino acid sequence of the alpha subunit. One cDNA clone (alpha 1) was isolated from a chicken embryo cDNA expression library in lambda gt11 by screening with anti-alpha-subunit polyclonal immunoglobulins. This alpha 1 cDNA contains an open reading frame of 1401 base pairs. A comparison of the translation of the nucleotide sequence with protein sequences obtained from the purified chicken alpha-subunit polypeptide verified that alpha 1 cDNA encoded the alpha subunit. Polymerase chain reactions were used to extend the sequence of alpha 1 cDNA toward the 5' end of alpha-subunit mRNA. The mature alpha subunit is composed of 516 amino acids with a calculated molecular mass of 59,373 Da. The compiled amino acid sequence contains two potential glycosylation sites, an observation that agrees with a previous demonstration that the alpha subunit contains two N-linked oligosaccharide chains. Blot hybridization analysis of total chicken embryo RNA detected an mRNA of 3.5 kilobases, a size that closely resembles the size of the cloned cDNA. Since the expression of the alpha subunit is confined to cell types that synthesize and secrete collagens, the regulation of the synthesis of the alpha subunit may play a central role in determining the expression of prolyl 4-hydroxylase activity.
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PMID:Prolyl 4-hydroxylase: molecular cloning and the primary structure of the alpha subunit from chicken embryo. 255 42

A single polypeptide acts as the beta subunit of prolyl 4-hydroxylase and the enzyme protein disulfide isomerase and may also function as a cellular thyroid hormone binding protein. We report here that the human gene for this polypeptide is about 18 kilobase pairs and consists of 11 exons. The two thioredoxin-like regions are coded by exons 1-2 and 8-9, respectively. The codons for the two presumed active sites of protein disulfide isomerase, each a Cys-Gly-His-Cys sequence, are located 12 base pairs from the beginning of exons 2 and 9. The last 3 amino acids coded by exons 1 and 8 and the first 9 amino acids coded by exons 2 and 9, including a broken codon for Tyr, are identical in the respective exon-intron junctions. These regions are also highly homologous to the active sites of bacterial thioredoxins. The data suggest that evolution of this gene has involved exon shuffling and duplication of a two-exon unit, in which the internal exon-intron junctions have been entirely conserved. The region between exons 1-2 and 8-9 appears to contain other duplications. The 5' flanking sequences contain a TATA box, six CCAAT boxes, and other elements which may be involved in regulation of the cellular amounts of this polypeptide.
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PMID:Characterization of the human gene for a polypeptide that acts both as the beta subunit of prolyl 4-hydroxylase and as protein disulfide isomerase. 284 39

Prolyl 4-hydroxylase was isolated in a highly purified form from a multi-cellular green alga, Volvox carteri, by a procedure consisting of ion-exchange chromatography and affinity chromatography on poly(L-hydroxyproline) coupled to Sepharose. Two other affinity-column procedures were also developed, one involving 3,4-dihydroxyphenylacetate and the other 3,4-dihydroxyphenylpropionate linked to Sepharose. The Km values of the Volvox enzyme for the co-substrates and the peptide substrate, as well as the inhibition constants for selected 2-oxoglutarate analogues, were similar to those of the enzyme from Chlamydomonas reinhardii, except that the Km for 2-oxoglutarate with the Volvox enzyme was 6-fold greater. The temperature optimum of the Volvox enzyme was also 10 degrees C higher. The apparent Mr of the Volvox enzyme by gel filtration was about 40,000, being similar to that reported for the Chlamydomonas enzyme but markedly lower than that of the vertebrate enzymes. A similar apparent Mr of about 40,000 was also found for prolyl 4-hydroxylase from the green alga Enteromorpha intestinalis, whereas the enzyme from various vascular plants gave an apparent Mr greater than 300,000. SDS/polyacrylamide-gel electrophoresis demonstrated in the highly purified Volvox enzyme the presence of a major protein band doublet with a Mr of about 65,000 and a minor doublet of Mr about 55,000-57,000. A polyclonal antiserum, prepared against the Mr-65,000 doublet, stained in immunoblotting the Mr-65,000 doublet as well as the alpha subunit, but not the beta subunit, of the vertebrate prolyl 4-hydroxylase. An antiserum against the beta subunit of the vertebrate enzyme stained in immunoblotting a Mr-50,000 polypeptide in a partially purified Volvox enzyme preparation, but did not stain either the Mr-65,000 or the Mr-55,000-57,000 doublet of the highly purified enzyme. The data thus suggest that the active Volvox carteri prolyl 4-hydroxylase is an enzyme monomer antigenically related to the alpha subunit of the vertebrate enzyme.
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PMID:Prolyl 4-hydroxylase from Volvox carteri. A low-Mr enzyme antigenically related to the alpha subunit of the vertebrate enzyme. 285 81

Several recent studies indicate that a single polypeptide may act as the beta-subunit of prolyl 4-hydroxylase, the enzyme protein disulphide-isomerase and a cellular thyroid-hormone-binding protein. We report here the isolation and characterization of cDNA clones encoding this multifunctional protein in the chicken. All the coding sequences were determined on the basis of nucleotide sequencing of five cDNA clones and amino acid sequencing of the N-terminal end of the chicken beta-subunit. The processed polypeptide contains 493 amino acid residues, the size of the respective mRNA being about 2.7 kb. The chicken beta-subunit cDNA sequences were 78% homologous to the previously reported human beta-subunit cDNA sequences at the nucleotide level and 85% homologous at the amino acid level. The homology of the chicken beta-subunit sequences to those reported for bovine thyroid-hormone-binding protein and rat protein disulphide-isomerase was also 85% at the amino acid level. Primary-structure comparisons between the four species indicated that the two proposed active sites of protein disulphide-isomerase, the two Trp-Cys-Gly-His-Cys-Lys sequences, are located within highly conserved regions, which are also homologous to the active sites of a number of thioredoxins. The middle of the polypeptide has an additional conserved region 100 amino acid residues in length in which the degree of homology between the four species is 94% at the amino acid level. This long conserved region may also be important for some of the multiple functions of the protein. The four extreme C-terminal amino acids of the polypeptide in all four species are Lys-Asp-Glu-Leu, a sequence that has been suggested to function as a signal for the retention of a protein in the endoplasmic reticulum.
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PMID:Molecular cloning of a multifunctional chicken protein acting as the prolyl 4-hydroxylase beta-subunit, protein disulphide-isomerase and a cellular thyroid-hormone-binding protein. Comparison of cDNA-deduced amino acid sequences with those in other species. 285 99

A single polypeptide is shown to act both as the beta subunit of the proline hydroxylase (EC 1.14.11.2) and as a protein disulfide-isomerase (EC 5.3.4.1). When isolated from chick embryos or rat liver, the beta subunit of prolyl 4-hydroxylase and the enzyme protein disulfide-isomerase have identical molecular weights and peptide maps as produced by digestion with Staphylococcus aureus V8 protease. The apparent molecular weights of both proteins isolated from human placental tissue are slightly higher, and the human beta subunit and one of its peptides have molecular weights about Mr 500 higher than the protein disulfide-isomerase and its corresponding peptide. Experiments with polyclonal and monoclonal antibodies also suggest a structural identity between the two proteins. The beta subunit isolated from the prolyl 4-hydroxylase tetramer has protein disulfide-isomerase activity similar to protein disulfide-isomerase itself, and even the beta subunit when present in the prolyl 4-hydroxylase tetramer has one-half of this activity.
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PMID:A single polypeptide acts both as the beta subunit of prolyl 4-hydroxylase and as a protein disulfide-isomerase. 303 69

Prolyl 4-hydroxylase (EC 1.14.11.2), an alpha 2 beta 2 tetramer, catalyses the formation of 4-hydroxyproline in collagens by the hydroxylation of proline residues in peptide linkages. We report here the isolation of cDNA clones coding for the beta-subunit of prolyl 4-hydroxylase from a human hepatoma lambda gt11 library and a corresponding human placenta library. Five overlapping clones covering all the coding sequences and almost all the non-coding sequences were characterized. The size of the mRNA hybridizing with these clones in Northern blotting is approximately 2.5 kb. The clones encode a polypeptide of 508 amino acid residues, including a signal peptide of 17 amino acids. These human sequences were found to be very similar to those recently reported for rat protein disulphide isomerase (EC 5.3.4.1). The degree of homology between these two proteins was 84% at the level of nucleotide sequences or 94% at the level of amino acid sequences. Southern blot analyses of human genomic DNA with a cDNA probe for the beta-subunit indicated the presence of only one gene containing these sequences. The product of a single gene thus appears to possess two different enzymatic functions depending on whether it is present in cells in monomer form or in the prolyl 4-hydroxylase tetramer.
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PMID:Molecular cloning of the beta-subunit of human prolyl 4-hydroxylase. This subunit and protein disulphide isomerase are products of the same gene. 303 2

Product inhibition of lysyl hydroxylase (peptidyllysine, 2-oxoglutarate:oxygen 5-oxidoreductase, EC 1.14.11.4) was studied with succinate, CO2, dehydroascorbate and hydroxylysine-rich polypeptide chains. The product inhibition patterns and addition data are consistent with a reaction mechanism involving an ordered binding of Fe2+, alpha-ketoglutarate, O2 and the peptide substrate to the enzyme in this order, and an ordered release of the hydroxylated peptide, CO2, succinate and Fe2+, in which Fe2+ need not leave the enzyme during each catalytic cycle and in which the order of release of the hydroxylated peptide and CO2 is uncertain. Ascorbate probably reacts by a substitution mechanism, either after the release of the hydroxylated peptide, CO2 and succinate or after the release of all products, including Fe2+, and dehydroascorbate is released before the binding of Fe2+. It is suggested that the ascorbate reaction is required to reduce either the enzyme-iron complex or the free enzyme, which may be oxidized by a side-reaction during some catalytic cycles, but not the majority. The mechanisms of the prolyl 4-hydroxylase and lysyl hydroxylase reactions are suggested to be identical. Zn2+, several citric acid cycle intermediates, nitroblue tetrazolium and homogentisic acid inhibited lysyl hydroxylase competitively with regard to Fe2+, alpha-ketoglutarate, O2 and ascorbate respectively, and epinephrine non-competitively with regard to all cosubstrates. Apparent Ki values are given for the product and other inhibitors.
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PMID:Studies on the lysyl hydroxylase reaction. II. Inhibition kinetics and the reaction mechanism. 676 67


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