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)

Bovine pancreatic deoxyribonuclease I (DNase I), an endonuclease that degrades double-stranded DNA in a nonspecific but sequence-dependent manner, has been used as a biochemical tool in various reactions, in particular as a probe for the structure of chromatin and for the helical periodicity of DNA on the nucleosome and in solution. Limited digestion by DNase I, termed DNase I 'footprinting', is routinely used to detect protected regions in DNA-protein complexes. Recently, we have solved the three-dimensional structure of this glycoprotein (relative molecular mass 30,400) by X-ray structure analysis at 2.5 A resolution and have subsequently refined it crystallographically at 2.0 A. Based on the refined structure and the binding of Ca2+-thymidine 3',5'-diphosphate (Ca-pTp) at the active site, we propose a mechanism of action and present a model for the interaction of DNase I with double-stranded DNA that involves the binding of an exposed loop region in the minor groove of B-DNA and electrostatic interactions of phosphates from both strands with arginine and lysine residues on either side of this loop. We explain DNase I cleavage patterns in terms of this model and discuss the consequences of the extended DNase I-DNA contact region for the interpretation of DNase I footprinting results.
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PMID:Structure of DNase I at 2.0 A resolution suggests a mechanism for binding to and cutting DNA. 371 45

From skin of Xenopus laevis, cDNA libraries were constructed and clones coding for the precursors of caerulein were isolated and sequenced. Using restriction endonuclease digestions, three different types of preprocaerulein cDNAs could be discerned. These were termed types I, III, and IV in accordance with the number of caerulein copies present in the sequence, the type III being the most abundant one. An incomplete copy of a fourth variant, termed type I', was also found. Besides deletions/insertions encompassing one or two caerulein sequences, these types also differ from each other by several point mutations. In the homologous precursor polypeptides deduced from the nucleotide sequence of these cloned cDNAs, the caerulein copies are flanked by complex processing sequences. These are Arg-Arg-Phe-Ala-Asp-Gly or Arg-Arg-Asp-Gly at the amino-terminal side and Gly-Arg-Arg at the carboxyl end. Between caerulein copies, highly homologous segments are present both at the polypeptide and cDNA level. This homology is evident both within a given precursor, where up to three such segments are present, and between the different types of precursors. We conclude that preprocaerulein cDNAs in the skin of X. laevis represent a small family, at least part of which is derived from different genes rather than being formed by alternative splicing of pre-mRNAs.
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PMID:Sequence of preprocaerulein cDNAs cloned from skin of Xenopus laevis. A small family of precursors containing one, three, or four copies of the final product. 375 78

ARGRII is one of the three regulatory genes controlling arginine metabolism in yeast. From a pool of hybrid plasmids carrying Sau3A fragments representing the entire yeast genome, a DNA fragment containing the regulatory gene ARGRII was cloned by complementation of an argRII- mutation, which prevents growth on ornithine as sole nitrogen source. Cells containing the cloned DNA regained the ability to repress the synthesis of anabolic enzymes and to induce the synthesis of the catabolic ones, when arginine is present. The 6.2 kb cloned DNA fragment encodes five transcripts (2.8 kb, 1.3 kb, 0.75 kb, 0.45 kb, 0.45 kb), which were located by S1 endonuclease mapping. By marker rescue the argRII- mutations were mapped in the DNA region coding for the 2.8 kb transcript, showing its importance in the control mechanism. Subcloning experiments confirm this result. However, at present the role of the 0.75 kb and 1.3 kb transcripts in the ARGR+ phenotype is unclear.
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PMID:Isolation and characterization of the yeast ARGRII gene involved in regulating both anabolism and catabolism of arginine. 388 75

The Eco RI endonuclease and methylase recognize the same hexanucleotide substrate sequence. We have determined the sequence of a fragment of DNA which encodes these enzymes using the chain-termination method of Sanger (Sanger, F., Nicklen, S., and Coulson, A. R. (1977) Proc. Natl. Acad. Sci. U. S. A. 74, 5463-5467). The amino acid sequences of both enzymes were derived from the DNA sequence. The coding regions selected include the only open translational frames of sufficient length to accommodate the enzymes. They coincide with previously established gene boundaries and orientation. The predicted amino acid sequences correlate well with analyses of the purified protein. Comparison of the nucleotide and protein sequences reveals no homology between the endonuclease and methylase which might provide insight into the origin of the restriction-modification system or the mechanism of common substrate recognition. Based on secondary structure predictions, the two enzymes also have grossly different molecular architecture. The base composition of the sequence is 65% A + T, and the codon usage is significantly different from that observed in several Escherichia coli chromosomal genes. In some cases, frequently selected codons are recognized by minor tRNA species. A spontaneous mutation in the endonuclease gene was isolated. Serine replaces arginine at residue 187. In crude extracts, Eco RI specific cleavage is approximately 0.3% wild type.
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PMID:Sequence analysis of the DNA encoding the Eco RI endonuclease and methylase. 625 3

We describe the construction and characterization of a cDNA plasmid for one of the rat liver glutathione S-transferase subunits. Poly(A)-RNA isolated from rat livers was enriched for glutathione S-transferase mRNA activity and used as templates to synthesize double stranded cDNA. The double stranded cDNAs were annealed to pBR322 through terminal deoxynucleotidyl transferase generated GC-tails followed by transformation into E. coli. Several candidate clones were selected by colony hybridization using polynucleotide kinase labeled liver and testis poly(A)-RNA probes. These candidate clones were further characterized by hybrid-selected translation of mRNA followed by immunoprecipitation and SDS gel electrophoresis. The positive clone, pGTR112 was mapped with restriction endonuclease analysis and sequenced by the chemical method of Maxam and Gilbert. The largest upen reading frame contains 142 amino acids very rich in Arg and Lys residues. The C-terminal residue phenylalanine of this open reading frame is consistent with what was reported for one of the ligandin subunits by Bhargava et al., (J. Biol. Chem. 253, 4116-4119, 1978). Among the 352 nucleotides covered by both pGTR112 and pGST94 described by Kalinyak and Taylor (J. Biol. Chem. 257, 523-530, 1982), there are only 9 nucleotide differences resulting in four changes of amino acid sequences.
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PMID:Cloning and sequence analysis of a cDNA plasmid for one of the rat liver glutathione S-transferase subunits. 629 39

The white pock variant of cowpox virus shows limited growth in chick embryo fibroblasts maintained in arginine-deprived culture medium. Since these conditions inhibit the growth of parental virus, there is a marked increase in the frequency of the white variant in the virus population recovered after passage in the absence of arginine. The variants generated in this system have been characterized by restriction endonuclease analysis of virus DNA in the total DNA recovered from infected cell cultures. Such analysis shows that the white variants arise as deletion mutants of the parental virus, but there was considerable heterogeneity in the restriction patterns of different isolates examined shortly after their generation. Further passage selected white cowpox virus populations with a stable genome configuration comparable with the DNA of pock-purified white variants.
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PMID:Arginine deprivation and the generation of white variants in cowpox virus-infected cell cultures. 629 24

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.
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PMID:Comparison of the cloned H-2Kbm1 variant gene with the H-2Kb gene shows a cluster of seven nucleotide differences. 630 Aug 87

The free energy of the binding reaction between EcoRI restriction endonuclease and a specific cognate dodecadeoxynucleotide (d(CGCGAATTCGCG)) has contributions from both electrostatic and nonelectrostatic components. These contributions were dissected by measuring the effects of varying salt concentration on the equilibrium binding constant and applying the thermodynamic analyses of Record et al. (Record, M. T., Jr., Lohman, T. M., and deHaseth, P. L. (1976) J. Mol. Biol. 107, 145-158). Endonuclease mutation S187 (Arg 187 to Ser) (Greene, P. J., Gupta, M., Boyer, H. W., Brown, W. E., and Rosenberg, J. M. (1981) J. Biol. Chem. 256, 2143-2153) did not significantly affect the nonelectrostatic component but did perturb the electrostatic contribution to the binding energy (we are numbering the amino acid residues according to the DNA sequence). The former was determined by extrapolating the linear portion of the salt dependence curve (0.125 to 0.25 M KCl) to 1 M ionic strength, with the same result for both wild type and S187 endonucleases at both pH 6.0 and 7.4 (-8.5 +/- 1.5 kcal/mol or greater than 50% of the total binding free energy). The slopes of these same curves yield estimates of eight ionic interactions between wild type endonuclease and the DNA at both pH values. By contrast, binding of EcoRI-S187 to dodecanucleotide involves six charge-charge interactions at pH 6.0. Only two ionic interactions are observed at pH 7.4. This was unexpected since gel permeation chromatography demonstrated that the recognition complex for both wild type and S187 proteins contains an enzyme dimer and a DNA duplex. EcoRI-S187 endonuclease retains wild type DNA sequence specificity, and the rate of the phosphodiester hydrolysis step is also unchanged. Thus, electrostatic interactions are functionally separable from sequence recognition and strand cleavage. Our results also establish that arginine 187 plays a key role in the electrostatic function and suggest that it might be located at the DNA-protein interface. The disproportionate loss of ion pairs at pH 7.4 can be rationalized by a model which suggests that six conformationally mobile ionic groups on the protein act in a coordinated manner during the interaction with DNA.
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PMID:Coordinate ion pair formation between EcoRI endonuclease and DNA. 631 32

BamHI methylase has been purified to apparent homogeneity. The isolated form of the enzyme is a single polypeptide with a molecular weight of 56,000 as determined by sodium dodecyl sulfate-polyacrylamide electrophoresis. Unlike BamHI endonuclease, which is isolated as a dimer and higher aggregates, the methylase has no apparent higher form. The methylase requires S-adenosyl-L-methionine as the methyl-group donor and is inhibited by Mg2+. The enzyme is also inhibited by 2,3-butanedione and reagents specific for sulfhydryl groups, such as N-ethylmaleimide, which suggests a role for arginine and cysteine residues, respectively. DNA efficiently protects the enzyme against the butanedione modification while S-adenosylmethionine has no effect. In contrast, S-adenosylmethionine protects against cysteine modification while DNA produces only small amounts of protection. Studies on the mechanism of methylation indicate that both strands of the recognition sequence are modified in a single binding event. The sequence specificity of the methylase is relaxed upon the addition of glycerol in the reaction mixture. In the presence of 30% glycerol the enzyme methylates sequences that are also recognized by BamHI endonuclease when acting under conditions of relaxed specificity.
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PMID:Sequence-specific BamHI methylase. Purification and characterization. 646 68

A Japanese family is described in which a 7-year-old child had isolated growth hormone deficiency type 1A, as described by Illig et al. He was shown to be homozygous for a deletion of the structural gene for hGH (hGH-N gene). Initially his growth rate responded well to hGH administration, but rapidly he developed high titers of hGH antibodies, and growth ceased. At that time, a somatomedin-C generation test gave negative results, suggesting that the growth arrest was related to the inability of hGH to generate somatomedin. Both parents were heterozygous for the hGH-N gene deletion and had a low hGH response to arginine and L-dopa tolerance tests, but had normal basal somatomedin-C levels and normal somatomedin-C generation tests. This family is the fourth to be reported with IGHD type 1A caused by deletion of the hGH-N gene. This cause of growth hormone deficiency can be distinguished from other severe autosomal recessive types of hGH deficiency by the demonstration of the deletion of hGH-N gene using restriction endonuclease analysis.
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PMID:Isolated growth hormone deficiency type 1A in a Japanese family. 653 75


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