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

---

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

A gene specifying tyrosine transfer RNA has been purified and transcribed in vitro. The purification procedure made use of two specialized transducing phages carrying the tRNA(Tyr) gene of Escherichia coli inserted into their DNA in opposite orientations. The separated heavy strands of the two phages were annealed and the single-stranded tails of the resulting hybrid were removed by digestion with Neurospora endonuclease. The size of the purified double-stranded structures was determined by electron microscopy. These isolated duplexes served as template for the in vitro transcription of tRNA(Tyr)-like molecules.
...
PMID:Purification and in vitro transcription of a transfer RNA gene. 494 95

We have carried out studies on type II restriction endonuclease EcoRI, which cleaves the DNA sequence 5'd(-G-A-A-T-T-C-)3', as indicated. The active form of the enzyme consists of two subunits, each 31063 molecular weight. A water-soluble reagent, 1-cyclohexyl-3-(2-morpholinoethyl)carbodiimide metho-p-sulphonate, which reacts with carboxyl groups and also with tyrosine and cysteine residues, has been found to inactivate this enzyme. Results are presented which show the following. (1) This specific inactivation is not due to modification of tyrosine or cysteine residues. (2) There is one carboxyl group per subunit which, when modified with carbodiimide, inactivates the enzyme. (3) phi X174 DNA (which does not contain EcoRI sites) partially protects the enzyme from the carbodiimide; protection is unaffected by the additional presence of Mg2+, but significantly greater with Co2+ and phi X174 DNA.
...
PMID:The essential carboxyl group in restriction endonuclease EcoRI. 627 65

The gene for the H-2K class I antigen of the bm1 variant was cloned and analyzed at the DNA level and compared with the previously cloned parent B6/Kh gene. Sequence determination and comparative restriction endonuclease studies indicate that Kbm1 is derived from the Kb gene. Seven nucleotide changes within a 13-nucleotide stretch distinguish the mutant from the parent gene and result in amino acid differences at positions 152, 155, and 156 in the antigen. The data confirm previously reported changes at amino acid positions 155 and 156 (arginine to tyrosine and leucine to tyrosine, respectively) and extend the altered region to include two nucleotides encoding a glutamate to alanine substitution at amino acid 152, a change not detected by the protein studies because of limitations of the methods used. The DNA sequence encoding this region of the Kbm1 glycoprotein is identical to the DNA sequence of at least one other known class I gene in the mouse, a finding consistent with the hypothesis that the mutation was not a random event but may be the result of a block transfer of information by a copy mechanism analogous to gene conversion. As the sequence analysis of the coding region for the first 273 amino acid residues shows identity between parent and mutant except for the seven nucleotide changes, all variant-parent functional differences must depend only on the cluster of three amino acid differences in the second domain of the Kb glycoprotein.
...
PMID:Comparison of the cloned H-2Kbm1 variant gene with the H-2Kb gene shows a cluster of seven nucleotide differences. 630 Aug 87

Modified polyomavirus genomes that individually encode the large and small T proteins were constructed by exchanging restriction endonuclease fragments between cDNA copies of the respective mRNAs and cloned genomic DNA. The efficacies of the new constructs, and that of the middle T protein gene described previously (R. Treisman , U. Novak, J. Favaloro , and R. Kamen , Nature [London] 292:595-600, 1981), were demonstrated with simian virus 40 (SV40)-polyomavirus recombinants in which part or all of the SV40 late region was replaced with the modified polyomavirus early genes. Each of the three recombinant viruses induced the synthesis of only the expected polyomavirus early protein in infected CV-1 cells. The rates of synthesis of large, middle, and small T proteins were ca. 1.5, 4.0, and 9.0 times the rate of synthesis of SV40 large T protein, respectively. The deletion of introns had no detrimental effect on mRNA biogenesis. Indeed, a further polyomavirus-SV40 recombinant, containing wild-type polyomavirus early region DNA, expressed an aberrant 58,000-dalton form of the middle T protein which we believe to result from utilization of a cryptic splice site. Immunofluorescence studied with monkey cells infected by the recombinant viruses allowed us to determine the cellular locations of the polyomavirus early proteins. Overproduction of the middle T protein did not result in a corresponding overproduction of the middle T protein-associated tyrosine phosphokinase activity.
...
PMID:Construction and functional characterization of polyomavirus genomes that separately encode the three early proteins. 632 36

Bisulfite mutagenesis techniques have been used to introduce single-point mutations within a region of the Rous sarcoma virus src gene defined by a BglI restriction endonuclease cleavage site. The mutants of Rous sarcoma virus that are produced by these techniques encode src proteins which contain single amino acid changes within a highly conserved amino acid sequence encompassing residues 430 to 433. DNA from the mutants CHpm26 ( Ala430 to Val), CHpm9 ( Pro431 to Ser), CHpm6 ( Glu432 to Lys), and CHpm65 ( Ala433 to Thr) each failed to transform chicken cells upon transfection, whereas DNA from CHpm59 (a third base alteration in the codon for Glu432 ) readily transformed chicken cells. Analysis of immune complexes containing the altered src proteins indicates that these proteins have decreased tyrosine protein kinase activity in vitro. In vivo labeling of cells infected with the mutant virus revealed diminished levels of the tyrosine-phosphorylated 34,000-molecular-weight protein. These data indicate that mutations within the sequence Ala430 - Pro431 - Glu432 - Ala433 lead to alterations in pp60src-specific tyrosine protein kinase activity and a concomitant loss of transforming potential of the mutant virus.
...
PMID:Amino acid alterations within a highly conserved region of the Rous sarcoma virus src gene product pp60src inactivate tyrosine protein kinase activity. 632 73

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.
...
PMID:Mutational analysis of a conserved motif of Agrobacterium tumefaciens VirD2. 747 69

Bovine pancreatic deoxyribonuclease I is an endonuclease of low specificity that interacts with the minor groove of DNA. Two amino acids, R41 and Y76, completely fill this groove, with R41 hydrogen bonding to the O2/N3 positions of pyrimidines and purines, and Y76 contacting a deoxyribose via an unusual hydrophobic "stacking" interaction. The roles of these amino acids in phosphodiester bond cleavage and in DNA hydrolysis selectivity have been studied by site-directed mutagenesis. Alterations have been made that are either conservative (R41K, Y76F) or more drastic (R41A, R41G, Y76A, Y76G). The surface loop (residues 73 to 76) that contains Y76 has also been deleted. Several double mutants in which both R41 and Y76 have been altered have also been prepared. The integrity of the catalytic site of the mutants has been investigated using the small, non-DNA, chromophoric substrate deoxythymidine-3',5'-di-(p-nitrophenyl)-phosphate. Hydrolysis of this compound was hardly changed, even by the most extreme alterations to R41 and Y76. In contrast, all the mutants bound DNA about ten times more weakly than the wild-type and, with the exception of R41K and Y76F, hydrolysed DNA much more slowly. This suggests that changes to R41 and Y76 have little effect on catalytic amino acids at the hydrolysis site, but are required to bind DNA and, more importantly, to correctly position the scissile phosphate for efficient hydrolysis. The selectivity of DNA hydrolysis for all the mutants has been tested using the 160 base-pair Escherichia coli Tyr T promoter DNA fragment. Very small differences were seen in global hydrolysis selectivity when either amino acid was altered. However, changes to R41 resulted in some differences to local cutting specificity that could be explained by the role of this amino acid in hydrogen bonding to particular bases relative to the scissile phosphate.
...
PMID:The roles of arginine 41 and tyrosine 76 in the coupling of DNA recognition to phosphodiester bond cleavage by DNase I: a study using site-directed mutagenesis. 765 Jul 37

Hemoglobin (Hb) M-Saskatoon, a beta variant of methemoglobin, is characterized by mild hemolysis. It is caused by the substitution of a histidine by a tyrosine at the 63rd amino acid residue of the beta-globin chain. Amplification and sequence analysis of genomic beta-globin DNA from an Indonesian boy diagnosed as having the more severe disease thalassemia demonstrated the presence of a C to T transition at nucleotide 473 in one of the two beta-globin genes resulting in a histidine to tyrosine substitution at 63rd residue. This amino acid change matched with that reported in Hb M-Saskatoon. This nucleotide change abolished a recognition site for the restriction endonuclease NlaIII. NlaIII digestion of the corresponding beta-globin DNA amplified from the patient's parents indicated that the mutation was inherited through from his mother. This result shows that the world-wide distribution of Hb M-Saskatoon extends to Indonesia, where it was not previously identified. Possible causes of the unusually severe symptoms observed in the case are discussed.
...
PMID:C to T transition at the first nucleotide of codon 63 of the beta-globin gene corresponding to hemoglobin M-Saskatoon in an Indonesian boy. 766

Negative selection of self-reactive immature T cells is mediated by TCR engagement and is thought to occur via apoptosis (programmed cell death). The requirement for the co-receptors CD4 and CD8 in negative selection has been demonstrated, but the biochemical mechanisms underlying their involvement in this process remain undefined. Here we present evidence that co-receptor engagement dramatically enhances CD3-induced endonuclease activation and cell death characteristic of apoptosis in immature thymocytes. The responses are associated with increased tyrosine phosphorylation of a number of cellular substrates, including the gamma isoform of phospholipase C, and with increased association of tyrosine phosphoproteins, including the protein tyrosine kinase p56lck, with the TCR complex. Co-receptor engagement also potentiated CD3-mediated Ca2+ increases via a mechanism dependent upon tyrosine kinase activation. Sustained Ca2+ availability was found to be necessary for endonuclease activation and apoptosis to occur. We suggest that CD4 and CD8 may participate in negative selection by enhancing TCR/CD3-induced tyrosine kinase activation and sustained Ca2+ increases that lead to endonuclease activation and apoptosis in self-reactive CD4+ CD8+ thymocytes.
...
PMID:Co-receptor (CD4/CD8) engagement enhances CD3-induced apoptosis in thymocytes. Implications for negative selection. 807 59

The introduction of small mutations instead of null alleles into the mouse genome has broad applications to the study of protein structure-function relationships and the creation of animal models of human genetic diseases. To test a simple mutational strategy we designed a targeting vector for the mouse proopiomelanocortin (POMC) gene containing a single nucleotide insertion that converts the initial tyrosine codon of beta-endorphin 1-31 to a premature translational termination codon and introduces a unique Hpal endonuclease restriction site. The targeting vector also contains a neo cassette immediately 3' to the last POMC exon and a herpes simplex virus thymidine kinase cassette to allow positive and negative selection. Homologous recombination occurred at a frequency of 1/30 clones of electroporated embryonic stem cells selected in G418 and gancyclovir. 10/11 clones identified initially by a polymerase chain reaction (PCR) strategy had the predicted structure without evidence of concatemer formation by Southern blot analysis. We used a combination of Hpa I digestion of PCR amplified fragments and direct nucleotide sequencing to further confirm that the point mutation was retained in 9/10 clones. The POMC gene was transcriptionally silent in embryonic stem cells and the targeted allele was not activated by the downstream phosphoglycerate kinase-1 promoter that transcribed the neo gene. Under the electroporation conditions used, we have demonstrated that a point mutation can be introduced with high efficiency and precision into the POMC gene using a replacement type vector containing a retained selectable marker without affecting expression of the allele in the embryonic stem cells. A similar strategy may be useful for a wide range of genes.
...
PMID:Introduction of a point mutation into the mouse genome by homologous recombination in embryonic stem cells using a replacement type vector with a selectable marker. 839 2


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>

---