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: EC:3.1.27.5 (RNase)
17,967 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Semiempirical potential energy calculations have been carried out to locate possible binding sites for purine nucleoside phosphates at the RNase S active center. The nucleobase of adenosine or 8-oxoadenosine was shown to be accommodated at the pyrimidine binding site of the enzyme provided the nucleoside must be "syn". The mutual orientation of 8-oxoadenosine carbonyl group and NH-group of Thr-45 suggests a formation of a corresponding hydrogen bond in the enzyme--nucleotide complex. The formation of the hydrogen bond was postulated earlier to be essential for specific recognition of the substrate by RNase. The results obtained are in good agreement with the postulate. The "leaving group" binding site at the RNase S active center was found to be nonspecific toward conformation (syn-anti) of purine nucleotides studied. In addition, optimal conformation of dinucleosidemonophosphate bound to the enzyme active site was estimated for Pyr-P-Pur substrates.
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PMID:[Theoretical conformational analysis of noncovalent complexes of purine nucleotides with ribonuclease]. 712 59

High levels of pancreatic ribonucleases are found in ruminants, species that have a ruminant-like digestion and several species with coecal digestion. Pancreatic ribonucleases from several independently evolved species with ruminant-like digestion were investigated to test a hypothesis that glycosylation of ribonucleases may have some function in species with coecal digestion and that glycosylation of the enzyme may not be advantageous for ruminants. Ribonucleases from the hippopotamus, two-toed sloth and three-toed sloth were isolated by extraction with sulfuric acid and affinity chromatography. Complete amino acid sequences were determined for the ribonucleases from the hippopotamus and two-toed sloth and a partial sequence for the enzyme from the three-toed sloth. The amino acids 75-78 of hippopotamus ribonuclease were positioned by homology with other artiodactyl ribonucleases. In hippopotamus ribonuclease a heterogeneity was found at position 37, half of the molecules containing glutamine acid the other half lysine. Hippopotamus ribonuclease differs less from pig and bovine ribonuclease than these differ from each other, because more ancestral characteristics have been retained. Although hippopotamus ribonuclease contains all four Asn-X-Ser/Thr sequences previously found to be glycosylation sites in one or more pancreatic ribonucleases, only the sequence Ans-Met-Thr (34-36) is glycosylated in the variant with glutamine at position 37, while the variant with lysine at this position is carbohydrate-free. Both sloth ribonucleases are completely glycosylated at the sequence Ans-Met-Thr (34-36) with a simple type of carbohydrate chain. The amino acid sequence of two-toed sloth ribonuclease shows some interesting coupled replacements.
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PMID:Pancreatic ribonucleases of mammals with ruminant-like digestion. Amino-acid sequences of hippopotamus and sloth ribonucleases. 743 54

A cDNA clone, predicted to encode a variant form of the type 1 fibroblast growth factor receptor (FGFR1) containing a dipeptide Val-Thr (VT) deletion at amino acid positions 423 and 424 located within the juxtamembrane region, was isolated from a Xenopus embryo (stage 8 blastula) library. Sequence analysis of genomic DNA encoding a portion of the FGFR1 juxtamembrane region demonstrated that this variant form arises from use of an alternative 5' splice donor site. RNase protection analysis revealed that both VT- and VT+ forms of the FGFR1 were expressed throughout embryonic development, the VT+ being the major form. Amino acid position 424 is located within a consensus sequence for phosphorylation by a number of Ser/Thr kinases. We demonstrate that a VT+ peptide was specifically phosphorylated by protein kinase C (PKC) in vitro, but not by protein kinase A (PKA). A VT- peptide, on the other hand, was not a substrate for either enzyme. Phosphorylation levels of in vitro synthesized FGFR-VT+ protein by PKC were twice that of FGFR-VT- protein. In a functional assay, Xenopus oocytes expressing FGFR-VT- or FGFR-VT+ protein were equally able to mobilize intracellular Ca2+ in response to basic fibroblast growth factor (bFGF). However, pretreatment with phorbol 12-myristate 13-acetate significantly reduced this mobilization in oocytes expressing FGFR-VT+ while having little effect on oocytes expressing FGFR-VT-. These findings demonstrate that alternative splicing of Val423-Thr424 generates isoforms which differ in their ability to be regulated by phosphorylation and thus represents an important mechanism for regulating FGFR activity.
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PMID:Cloning of a fibroblast growth factor receptor 1 splice variant from Xenopus embryos that lacks a protein kinase C site important for the regulation of receptor activity. 755 2

Peptide-water interactions of a ribonuclease C-peptide analogue, RN-24 (Suc-AETAAAKFLRAHANH2), which exhibits significant helicity, have been studied in solution using homonuclear 2D and 3D NMR cross-relaxation experiments. Dipolar peptide proton-water proton interactions are indicated by a large number of NOESY-type cross peaks at the H2O resonance frequency, most of them with opposite sign relative to the diagonal. Some cross peaks arise from intrapeptide cross relaxation to labile protons of histidine, threonine, lysine and arginine side chains. The observed peptide-water interactions are rather uniformly distributed, involving peptide backbone and side chains equally. The data are consistent with rapid fluctuations of the conformational ensemble and the absence of peptide regions that are highly shielded from bulk solvent, even in a peptide that exhibits high propensities for formation of helical secondary structure.
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PMID:Hydration of the partially folded peptide RN-24 studied by multidimensional NMR. 764 54

Transforming growth factor-beta 1 (TGF-beta 1) induces angiogenesis in vivo and capillary morphogenesis in vitro. Two receptor serine/threonine kinases (types I and II) have been identified as signal transducing TGF-beta receptors. We explored the possibility of inhibiting TGF-beta-mediated events in glomerular capillary endothelial cells using a TGF-beta type II receptor (T beta R-II) transdominant negative mutant. A mutant TGF-beta type II receptor (T beta R-IIM), lacking the cytoplasmic serine/threonine kinase domain, was produced by polymerase chain reaction using rat T beta R-II cDNA as template. Since T beta R-II and TGF-beta type I receptor (T beta R-I) heterodimerize for signal transduction, the mutant receptor competes for binding to wild-type T beta R-I, hence acting in a dominant negative fashion. Glomerular capillary endothelial cells were stably transfected with T beta R-IIM, and four independent clones were expanded. That the T beta R-IIM mRNA was expressed was shown by reverse transcriptase-polymerase chain reaction, RNase protection assay, and Northern analysis. Presence of cell surface T beta R-IIM protein was shown by affinity cross-linking with 125I-TGF-beta 1. In wild-type endothelial cells, TGF-beta 1 (2 ng/ml) significantly inhibited [3H]thymidine incorporation to 63 +/- 10% of control (n = 4). In transfected endothelial cells carrying T beta R-IIM, TGF-beta 1 stimulated [3H]thymidine incorporation to 131 +/- 9% of control (n = 4, p < 0.005). Also, in wild-type endothelial cells, endogenous and exogenous TGF-beta 1 induced apoptosis and associated capillary formation. Both apoptosis and capillary formation were uniformly and entirely absent in transfected endothelial cells carrying T beta R-IIM. This represents the first demonstration that capillary morphogenesis in vitro is associated with apoptosis, and that interference with T beta R-II signaling inhibits this process in glomerular capillary endothelial cells.
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PMID:Inhibition of capillary morphogenesis and associated apoptosis by dominant negative mutant transforming growth factor-beta receptors. 767 46

The raf proto-oncogenes encode cytoplasmic protein serine/threonine kinases, which play a critical role in cell growth and development. One of these, A-raf-1 (human gene symbol, ARAF1), which is predominantly expressed in mouse urogenital tissues, has been mapped to an evolutionarily conserved linkage group composed of ARAF1, SYN1, TIMP, and properdin located at human chromosome Xp11.2. We have isolated human genomic DNA clones containing the expressed gene (ARAF1) on the X chromosome and a pseudogene (ARAF2) on chromosome 7p12-q11.21. Analysis of the nucleotide sequence from the ARAF1 genomic clones demonstrated that it consists of 16 exons encoded by minimally 10,776 nucleotides. The major transcriptional start site (+1) was determined by RNase protection and primer extension assays. Promoter activity was confirmed by functional assays using DNA fragments fused to a CAT reporter gene. The ARAF1 minimal promoter, located between nucleotides -59 and +93, has a low G + C content and lacks consensus TATA and Inr sequences but shows sequence similarity at position -1 to the E box that is known to interact with USF and TFII-I transcription factors.
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PMID:The complete sequence and promoter activity of the human A-raf-1 gene (ARAF1). 802 Sep 55

Different molecular forms of ribonuclease were isolated from fresh human pancreas obtained from healthy transplant donors. The purification procedure consists of the preparation of an acetone powder, followed by (NH4)2SO4 precipitation and two chromatography steps (cationic exchange and reversed-phase). Protein bands in gel electrophoresis with RNAase activity were monitored using a negative-staining zymogram technique. Several glycosylated enzyme forms with apparent molecular masses ranging from 14 to 40 kDa were separated. Peptides containing the three Asn-Xaa-Thr/Ser acceptor sites for glycosylation were isolated and analysed. The site with Asn-34 was almost completely glycosylated, while the sites with Asn-76 and Asn-88 had carbohydrate in about half and a minor part of the molecules, respectively. The carbohydrate compositions of the glycopeptides are different from those of the same gene product isolated from human urine. C-Terminal threonine was present in part of the molecules, indicating partial degradation by carboxypeptidase.
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PMID:Heterogeneity in the glycosylation pattern of human pancreatic ribonuclease. 807 10

The molecular basis for the enzymatic specificity of human angiogenin has been investigated by site-directed mutagenesis of Thr-44, Glu-108, and Ser-118--residues corresponding to those thought to be involved in substrate base recognition in the homologous protein, RNase A. Mutations of Thr-44 to Ala, His, and Asp affect both activity and specificity dramatically. The Ala and His replacements decrease activity toward tRNA by factors of 25 and 40, respectively, and reduce cleavage of cytidylyl more than uridylyl dinucleotides. Substitution by Asp does not influence the rate of tRNA and rRNA degradation but alters specificity even more markedly than the other mutations: T44D-angiogenin has 17-40-fold decreased activity toward CpN' dinucleotides and 1.3-1.9-fold increased activity toward UpN', resulting in an inverted order of preference (U > C) compared to native angiogenin. Mutations of Glu-108 to Lys and Gln change activity toward RNA and dinucleotides by no more than 50% and produce slight increases in preference for adenosine vs guanosine at position N' of NpN' substrates. Mutations of Ser-118 to Asp and Arg have a larger effect, decreasing activity by factors of approximately 2 and 4, respectively, toward all substrates examined. These results indicate that: (i) Thr-44 is important for recognition of the pyrimidine moiety at position N, (ii) Glu-108 may make a small contribution to binding the N'-nucleotide, and (iii) Ser-118 has a minor functional role, which appears to involve catalysis rather than nucleotide binding.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Alteration of the enzymatic specificity of human angiogenin by site-directed mutagenesis. 809 59

Serine-threonine protein kinases in the ribosomal S6 kinase (rsk or p90rsk) family have been implicated as signaling intermediates in the cellular response to several growth factors. To investigate the molecular diversity of human p90rsk isoforms, mixed degenerate oligonucleotide polymerase chain reaction was used to isolate partial rsk cDNAs (1.1 kb). Three closely related human rsk cDNAs were obtained (HU-1, HU-2, HU-3). These cDNAs are encoded by separate genes based on DNA sequence diversity and distinct patterns seen with genomic Southern blots. Northern analysis revealed different sized mRNA transcripts for each isoform. A full-length HU-1 cDNA (3.1 kb) was subsequently isolated from a HeLa cell library. 5'-cDNA clones for HU-2 and HU-3 were isolated using the "rapid amplification of cDNA ends" strategy. Experiments using human x hamster somatic cell hybrids localized the HU-1 gene to human chromosome 3; HU-2 is on chromosome 6; and HU-3 is on the X chromosome. The tissue distribution of human rsk mRNAs was determined using ribonuclease protection assays. HU-3 mRNA was present in multiple RNA samples. HU-2 was expressed in fibroblast > muscle > lymphocyte = placenta > liver. HU-1 was expressed in Epstein-Barr virus lymphocyte > > muscle = liver > fat = placenta. These results indicate that the multiplicity of p90rsk isoforms is increased to at least three for humans and that marked tissue-/cell-specific differences in p90rsk isoform expression are present.
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PMID:Human rsk isoforms: cloning and characterization of tissue-specific expression. 814 Dec 49

The adenomatous polyposis coli (APC) gene, responsible for familial adenomatous polyposis, is also associated with development of sporadic tumors in digestive system as colon, stomach, or pancreas. In order to investigate whether or not APC mutations occur as an early genetic event during gastric carcinogenesis, we examined somatic mutations of APC in flat adenomas of the stomach. DNAs isolated from flat adenomas were examined by means of an RNase protection analysis coupled with polymerase chain reaction (PCR) followed by DNA sequencing of the PCR products. By screening a mutation cluster region (MCR: codons between 1286 and 1513) of APC in which two-thirds of somatic mutations were detected in colorectal tumors, somatic mutations were found in four of ten flat adenomas: three of which caused truncation of the gene product due to a nonsense mutation or 4-bp deletion; one other was a point mutation that altered amino acid from alanine to threonine. Our results imply that APC plays a crucial role in an early step of gastric carcinogenesis, as was observed in colorectal carcinogenesis.
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PMID:Somatic mutations of the APC gene in precancerous lesion of the stomach. 824 71


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