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:3.1.30.2 (endonuclease)
18,621 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The hydrolysis of several tRNAs by an endonuclease extracted from the venom of Naja oxiana and specific for double-stranded, or at least highly ordered, regions has been studied under various experimental conditions. It is shown that the hydrolysis patterns of yeast tRNAPhe, tRNAVal and tRNAAsp in the isolated state are similar, most of the cuts occurring in the anticodon and acceptor stems. Ionic conditions are able to modify the hydrolysis pattern. The origin of these modifications is discussed. The protection against ribonuclease action, afforded to tRNAPhe, tRNAVal and tRNAAsp by the cognate aminoacyl-tRNA synthetase, is analyzed. It is shown that in all cases the anticodon stem is protected. The 3'-terminal region does not seem to be tightly engaged in the complex with the aminoacyl-tRNA synthetase. These results are discussed in the light of information on contact areas previously obtained by ultraviolet cross-linking techniques. The effects of the small ligands (ATP and amino acid) on the protection afforded to the tRNA by the cognate synthetase, have been studied. In the valine and aspartic acid systems, ATP induced a modification of the tRNA-enzyme complex leading to differences in the hydrolysis pattern of the 3'-accepting region. The effects of aminoacylation on the cleavage of tRNAPhe, tRNAVal and tRNAAsp were also studied. Whereas no modification of the cleavage map was observed in the aspartic system, aminoacylation resulted in slight but significant modifications of the hydrolysis pattern for tRNAPhe and tRNaVal in the 3'-terminal region.
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PMID:Comparison of the hydrolysis patterns of several tRNAs by cobra venom ribonuclease in different steps of the aminoacylation reaction. 691 54

The VirD2 polypeptide from Agrobacterium tumefaciens, in the presence of VirD1, introduces a site- and strand-specific nick at the T-DNA borders. A similar reaction at the origin of transfer (oriT) of plasmids is essential for plasmid transfer by bacterial conjugation. A comparison of protein sequences of VirD2 and its functional homologs in bacterial conjugation and in rolling circle replication revealed that they share a conserved 14 residue segment, HxDxxx(P/u)HuHuuux [residues 126-139 of VirD2; Ilyina, T.V. and Koonin, E.V. (1992) Nucleic Acids Res. 20, 3279-3285]. A mutational approach was used to test the role of these residues in the endonuclease activity of VirD2. The results demonstrated that the two invariant histidine residues (H133 and H135) are essential for activity. Mutations at three sites, histidine 126, aspartic acid 128 and aspartic acid 130, that are conserved in a subfamily of the plasmid mobilization proteins, led to the loss of VirD2 activity. Aspartic acid at position 130, could be substituted with glutamic acid and to a much lesser extent, with tyrosine. In contrast, another conserved residue, asparagine 139, tolerated many different amino acid substitutions. The non-conserved residues, arginine 129, proline 132 and leucine 134, were also found to be important for function. Isolation of null mutations that map throughout this conserved domain confirm the hypothesis that this region is essential for function.
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PMID:Mutational analysis of a conserved motif of Agrobacterium tumefaciens VirD2. 747 69

Escherichia coli RuvC protein is a specific endonuclease that resolves recombination intermediates into viable products. The structural features needed for RuvC activity were investigated by sequencing three ruvC mutations and relating the base pair changes identified to the activity of the mutant proteins. Each of the three mutations is a single base-pair substitution. ruvC51 converts glycine-15 to an aspartic acid residue. The product of ruvC51 was purified and shown to retain the ability to bind junctions, albeit with a slightly reduced affinity. However, it has lost the ability to resolve these structures by symmetrical cleavage. A multicopy ruvC51 plasmid confers sensitivity to UV light in a ruvC+ strain. The ruvC53 allele causes a glycine-17 to serine substitution while ruvC55 produces a stop codon. Neither of these genes produces a stable product. The results suggest that the N-terminal domain of RuvC may be concerned with cleavage of junctions.
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PMID:An E. coli RuvC mutant defective in cleavage of synthetic Holliday junctions. 839 86

We report a rare apolipoprotein E variant in an Irish female with Type III hyperlipidaemia who has the phenotype E2E1 as determined by isoelectric focusing. Sequence analysis of the apolipoprotein E gene from the proband and from four other family members, using DNA amplified by the polymerase chain reaction, demonstrated the presence of a point mutation in the common epsilon 2 allele with a G-->A transition at nucleotide 3791. This was confirmed by digestion with the restriction endonuclease TaqI, which cuts at a new site within the apolipoprotein E gene, created by the base change. This mutation results in a substitution of aspartic acid for glycine at position 127 of the mature protein. We believe this to be the first description of this apolipoprotein E variant in a family from the British Isles. The mutation appears to be 'recessive' with respect to the expression of Type III hyperlipidaemia, although it may be somewhat more potent in this regard than the parent epsilon 2 allele. The Type III hyperlipidaemia is responsive to treatment with diet and gemfibrozil.
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PMID:Rare apolipoprotein E variant identified in a patient with type III hyperlipidaemia. 850 53

Three novel point mutations were detected in the glucocerebrosidase gene of three unrelated Gaucher's disease patients by direct sequencing of PCR products. The first is a C to G change at position 4263 in the genomic sequence (exon 7) which results in a proline to arginine change at position 266 in the mature enzyme (P266R). The second is a G to C change at position 5276 in the genomic sequence (exon 8) which results in an aspartic acid to histidine change at position 315 (D315H). The third is a C to A change at position 5286 in the genomic sequence (exon 8) which results in an alanine to aspartic acid change at position 318 (A318D). The first mutation destroys an AvaII restriction endonuclease site, the second creates a BspMI site and the third creates a BamH I site.
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PMID:Three unrelated Gaucher's disease patients with three novel point mutations in the glucocerebrosidase gene (P266R, D315H and A318D). 854 70

The phenotypic characteristics of isolated growth hormone deficiency (IGHD) type IB in humans, such as autosomal recessive inheritance, time of onset of growth retardation, diminished secretion of growth hormone (GH) and IGF-I, proportional reduction in weight and size, and delay in sexual maturation, has much in common with the phenotype of the homozygous little/little (lit/lit) mouse. Sequencing of the GH releasing hormone (GHRH) receptor in lit/lit mice has shown a single nucleotide substitution within the extracellular peptide binding domain at codon 60 that changed aspartic acid to glycine. Therefore, the GHRH receptor is a reasonable candidate gene for causing IGHD in humans. DNA from 65 unrelated healthy Caucasians of normal stature and 65 children with IGHD type IB of whom 12 did not respond to exogenous treatment with GHRH were studied. Restriction endonuclease analysis, linkage studies, and polymerase chain reaction amplification and sequencing of the whole extracellular domain including the first three membrane spanning domains of the GHRH receptor gene were performed. None of the analyses revealed any structural abnormalities in these patients with IGHD. This suggests that a lit/lit mouse equivalent is an unlikely explanation for the majority of children with IGHD. Although gross structural abnormalities in the whole gene have been ruled out in this study, mutations in the carboxyl terminus are still possible, and, therefore, the remaining part of the gene needs to be sequenced.
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PMID:Isolated growth hormone deficiency: testing the little mouse hypothesis in man and exclusion of mutations within the extracellular domain of the growth hormone-releasing hormone receptor. 861 1

Phosphatidylcholine-specific phospholipase D (PLD) enzymes catalyze hydrolysis of phospholipid phosphodiester bonds, and also transphosphatidylation of phospholipids to acceptor alcohols. Bacterial and plant PLD enzymes have not been shown previously to be homologues or to be homologous to any other protein. Here we show, using sequence analysis methods, that bacterial and plant PLDs show significant sequence similarities both to each other, and to two other classes of phospholipid-specific enzymes, bacterial cardiolipin synthases, and eukaryotic and bacterial phosphatidylserine synthases, indicating that these enzymes form an homologous family. This family is suggested also to include two Poxviridae proteins of unknown function (p37K and protein K4), a bacterial endonuclease (nuc), an Escherichia coli putative protein (o338) containing an N-terminal domain showing similarities with helicase motifs V and VI, and a Synechocystis sp. putative protein with a C-terminal domain likely to possess a DNA-binding function. Surprisingly, four regions of sequence similarity that occur once in nuc and o338, appear twice in all other homologues, indicating that the latter molecules are bi-lobed, having evolved from an ancestor or ancestors that underwent a gene duplication and fusion event. It is suggested that, for each of these enzymes, conserved histidine, lysine, aspartic acid, and/or asparagine residues may be involved in a two-step ping pong mechanism involving an enzyme-substrate intermediate.
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PMID:A novel family of phospholipase D homologues that includes phospholipid synthases and putative endonucleases: identification of duplicated repeats and potential active site residues. 873 63

Hepatic angiosarcoma (HA) is an uncommon neoplasm associated with known etiologic factors in 25% to 42% of cases. It is, however, one of the most common sarcomas found in the liver. The aim of this study was to find was to find mutations in the K-ras-2 oncogene in sporadic and Thorotrast (TT)-induced HA. Point mutations in K-ras-2 were sought in archival, formalin-fixed tissue blocks from 24 patients with angiosarcoma. Of these, 19 cases were sporadic and 5 were TT-induced. Mutational analysis was performed by topographic microdissection with PCR amplification followed by genotyping. Specific mutations were determined by two independent methods: (a) direct sequencing of the PCR product confirmed by rePCR and by using a different sequencing primer, and (b) PCR-based selective enrichment of mutant DNA by endonuclease digestion followed by heteroduplex DNA analysis using denaturing gradient gel electrophoresis. Eleven K-ras-2 point mutations were detected in 7 of 24 (29%) tumors, including 5 of 19 (26%) sporadic HA and 2 of 5 (40%) TT-induced HA. There were seven G:C > A:T and four G:C > T:A mutations. All seven mutated tumors contained a codon 12-aspartate amino acid substitution. In addition, a second codon 12-cysteine mutant cell population was present in one of two codon 12-aspartate mutated TT-induced HA and in three of five codon 12-aspartate sporadic tumors. Of these four tumors, three contained both aspartate and cysteine mutations and were composed of multiple nodules; the fourth was a single mass. Seventeen tumors had multiple nodules; whereas 5 had a K-ras-2 mutation, 12 were wild-type. The molecular pathology of both sporadic and TT-induced HA is characterized by a high rate of K-ras-2 mutations characteristic of oxidative damage (ie, G:C > A:T and G:C > T:A mutations) resulting in two mutated population sets: codon 12 GGT > GAT and GGT > TGT (glycine to aspartic acid and cysteine). This is, to date, the first study to characterize the K-ras-2 gene mutations within human sporadic and TT-induced HA by direct sequence analysis and denaturing gradient gel electrophoresis. These data further support the hypothesis linking adduct-forming vinyl chloride exposure to HA containing a much higher frequency of K-ras-2 mutations and a mutational spectrum characteristic of chloroethylene oxide, a carcinogenic metabolite of vinyl chloride.
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PMID:Sporadic and Thorotrast-induced angiosarcomas of the liver manifest frequent and multiple point mutations in K-ras-2. 901 Apr 58

Superposition of the PI-SceI and I-CreI homing endonuclease three-dimensional x-ray structures indicates general similarity between the I-CreI homodimer and the PI-SceI endonuclease domain. Saddle-shaped structures are present in each protein that are proposed to bind DNA. At the putative endonucleolytic active sites, the superposition reveals that two lysine (Lys-301 and Lys-403 in PI-SceI and Lys-98 and Lys-98' in I-CreI) and two aspartic acid residues (Asp-218 and Asp-326 in PI-SceI and Asp-20 and Asp-20' in I-CreI) are related by 2-fold symmetry. The critical role of Lys-301, Asp-218, and Asp-326 in the PI-SceI reaction pathway was reported previously. Here, we demonstrate the significance of the active-site symmetry by showing that alanine substitution at Lys-403 reduces cleavage activity by greater than 50-fold but has little effect on the DNA binding activity of the mutant enzyme. Substitution of Lys-403 with arginine, which maintains the positive charge, has only a modest effect on activity. Interestingly, even though the Lys-301 and Lys-403 residues display pseudosymmetry, PI-SceI mutant proteins with substitutions at these positions have different behaviors. The presence of similar basic and acidic residues in many LAGLIDADG homing endonucleases suggests that these enzymes use a common reaction mechanism to cleave double-stranded DNA.
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PMID:Identification of Lys-403 in the PI-SceI homing endonuclease as part of a symmetric catalytic center. 980 21

Terminase, the DNA packaging enzyme of bacteriophage lambda, is a heteromultimer of gpNu1 and gpA subunits. In an earlier investigation, a lethal mutation changing gpA residue 497 from lysine to aspartic acid (K497D) was found to cause a mild change in the high-affinity ATPase that resides in gpA and a severe defect in the endonuclease activity of terminase. The K497D terminase efficiently sponsored packaging of mature lambda DNA into proheads. In the present work, K497D terminase was found to have a severe defect in the cohesive end separation, or helicase, activity. Plaque-forming pseudorevertants of lambda A K497D were found to carry mutations in A that suppressed the lethality of the A K497D mutation. The two suppressor mutations identified, A E515G and A E515K, affected residue 515, which is located near the putative P-loop of gpA. A codon substitution study of codon 515 showed that hydrophobic and basic residues suppress the K497D defect, but hydrophilic and acidic residues do not. The E515G change was demonstrated to reverse the endonuclease and helicase defects caused by the K497D change. Moreover, the gpA K497D E515G enzyme was found to have kinetic constants for the high-affinity ATPase center similar to those of the wild type enzyme, and the endonuclease activity of the K497D E515G enzyme was stimulated by ATP to an extent similar to the ATP stimulation of the endonuclease activity of the wild type enzyme.
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PMID:Endonuclease and helicase activities of bacteriophage lambda terminase: changing nearby residue 515 restores activity to the gpA K497D mutant enzyme. 1106 51


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