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)

A scheme for the isolation of Ca,Mg-dependent endonuclease from human spleen lymphocyte nuclei has been developed. The isolation procedure resulted in protein preparations (Mr = 57 kD) possessing an enzymatic activity and stable upon storage for over a period of one year. The enzyme is an endonuclease which predominantly cleaves double-stranded DNA by a mixed single- and double-hit mechanism with the formation of 5'-phosphate and 3'-OH terminal groups. Its maximal activation is induced by Ca2+ plus Mg2+. The enzyme is also active in the presence of Mn2+, Ca2+, Mg2+ and Zn2+ and is inhibited by Co2+. NaCl and KCl (0.15-0.2 M) and p-chloromercuribenzoate (1 mM) also inhibit the enzyme. ATP has no activating effect.
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PMID:[Isolation and analysis of Ca/Mg-dependent endonuclease from cell nuclei of human splenocytes]. 317 64

An endonuclease specific for cruciform junctions has been purified from yeast cells treated with a DNA-damaging agent. The activity was followed through five chromatographic steps by assaying for the linearization of supercoiled plasmid DNA, which extrudes cruciform structures in vitro. The sites of cleavage on pColIR215 were sequenced, and nicks were located to positions symmetrically opposed across the cruciform junction. The products of cleavage were unit length linear duplexes that contained terminal hairpin loops. In contrast to pColIR215, the cleavage patterns of pXG540 plasmid DNA were found to be complex, and cuts were found up to 40 bases from an (A-T)34 sequence that extrudes into a cruciform. Little or no activity could be detected on single-stranded DNA, linear duplex DNA, or nicked circular duplex DNA. The nuclease was insensitive to RNase but was inactivated by treatment with proteinase K. Mg2+ was required as cofactor and could not be replaced by Mn2+, Ca2+, Co2+, or Cu2+. The native molecular weight of the activity was approximately 200,000 as estimated by gel filtration.
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PMID:Purification and properties of a nuclease from Saccharomyces cerevisiae that cleaves DNA at cruciform junctions. 330 13

With the use of the strain-overproducer restriction endonuclease R.EcoRV was isolated and purified to homogeneity. The molecular mass of the enzyme was determined by gel filtration and polyacrylamide gel electrophoresis to be 25 000 daltons. According to the data of immunological tests R.EcoRV differs in its antigenic characteristics from restriction endonucleases R.EcoRI and R.EcoRII. Dependence of enzyme activity on pH, ionic strength, temperature, presence of divalent cations (Mn2+, Mg2+, Co2+, Zn2+, Ni2+ and Cd2+) and organic solvents (glycerol, dimethylsulfoxide, ethanol) has been studied. It was shown that under conditions of replacement of Mg2+ for Mn2+ or after addition of organic solvents relaxation of R.EcoRV specificity takes place. It was shown also that R.EcoRV is able to digest T-even bacteriophage DNAs with different types and extents of modification. DNA modified by the action of MR.EcoRV system in vivo is susceptible to R.EcoRV in vitro. Under conditions of relaxed specificity noncanonical sites are susceptible to R.EcoRV attack. The fragments resulted may be cloned in canonical pBR322 EcoRV site.
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PMID:[EcoRV restrictase: physical and catalytic properties of homogenous enzyme]. 620 Jul 65

An endodeoxyribonuclease has been purified from nuclei of bovine small intestinal mucosa to a homogeneous state by a procedure involving affinity chromatography on heparin-agarose. The endonuclease, which was found to be bound to chromatin, has a pH optimum of 5.4. It requires Mn2+ or Co2+ for activity and its maximum activity with Mg2+ is about 80% of that with Mn2+. Its activity is strongly inhibited by sulfhydryl-blocking agents, and by ethidium bromide. The enzyme does not attack RNA and is inhibited by it. Its isoelectric point is 8.5 +/- 0.1, and its molecular weight is 49,000 +/- 3,000, determined by sucrose gradient sedimentation and gel filtration on Sephadex G-100. Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate indicated that the enzyme is composed of two nonidentical subunits with molecular weights of 30,000 and 23,000. The enzyme catalyzes the endonucleolytic cleavage of circular duplex ColE1 DNA via single strand scissions from the initial stage of degradation. The average size of the limit products of native phage T7 or ColE1 DNA is about 2,000 to 1,500 base pairs, estimated by neutral sucrose gradient sedimentation or agarose gel electrophoresis. The enzyme degrades denatured DNA about 20 times faster than native DNA. The products contain 5'-phosphoryl and 3'-hydroxyl termini, and all four deoxymononucleotides are present in almost equal amounts at the 5'-termini.
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PMID:Purification and properties of an endodeoxyribonuclease from nuclei of bovine small intestinal mucosa. 625 82

Restriction endonuclease EcoRI cleaves the DNA sequence 5'd(-G-A-A-T-T-C-) under optimum digestion conditions. A variation in pH and ionic strength can result in EcoRI activity when 5'd(-A-A-T-T-) is cut. A divalent cation, usually Mg2+, is required for enzyme activity, though Mn2+ can also be used. Eight different cations with ionic radius/charge ratios similar to Mg2+ were tested and Co2+ and Zn2+ were also found to act as cofactors for EcoRI. A comprehensive study has been made of the effect of NaCl and pH on the EcoRI/EcoRI transition in the presence of the above four cations. Generally, a decrease in NaCl and/or an increase in pH caused a decrease in enzyme specificity. The changeover depended on the cation. They may be placed in order of their ability to increase EcoRI specificity thus: Co2+ greater than Zn2+ greater than Mg2+ greater than Mn2+. The Km of EcoRI for ColE1 DNA, in the presence of Co2+, was found to be 0.4 nM, compared to 3 nM with Mg2+, whereas the turnover was only one double-stranded scission/min with Co2+ compared to eight/min with Mg2+. The implications of all these findings on the enzyme's mechanism are discussed.
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PMID:Cation dependence of restriction endonuclease EcoRI activity. 626 25

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.
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PMID:The essential carboxyl group in restriction endonuclease EcoRI. 627 65

A specific type-II restriction endonuclease BcnI from Bacillus centrosporus has been purified to electrophoretic homogeneity in three chromatographic steps. Around 15 micrograms of such a preparation can be isolated from 1 g of the cell paste. The yield of the enzyme is higher than that of any type-II restriction endonuclease so far reported. The molecular weight of the enzyme determined by gel filtration and polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulphate equals 27 500 and 28 000 respectively. The activity of the restriction endonuclease is maximal at pH 9.2 and 40--45 degrees C. The optimal magnesium concentration was estimated to be 7.5 mM. The activity of BcnI may also be observed in the presence of Co2+, Mn2+, Ni2+ and Zn2+ but it is markedly less than in the presence of Mg2+.
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PMID:Isolation and some properties of the restriction endonuclease BcnI from Bacillus centrosporus. 627 27

An endodeoxyribonuclease has been purified to near homogeneity from rat small intestinal mucosa by a procedure involving Con A-Sepharose affinity chromatography. During the initial steps of purification, the presence of 5 mM CaCl2 was essential for stability of the enzyme activity. The enzyme has a molecular weight of 32 000 and an isoelectric point of 4.7. NaCl, sulfhydryl reagents, and iodoacetate strongly inhibited the reaction, but tRNA did not. The enzyme required divalent cations for activity and had a pH optimum of pH 6.2 with Co2+ and pH 7.7 with Mn2+. In both optimum conditions, the enzyme hydrolyzed native DNA more rapidly than denatured DNA, and the average chain lengths of limit digestion products of native and denatured DNA were 8 and 10, respectively, at pH 6.2 and 9 and 11, respectively, at pH 7.7. The enzyme activity to produce acid-soluble fractions from linear DNA substrate was similar in the two optimum conditions, but the activity to nick double-stranded, superhelical circular DNA substrate was significantly higher at pH 6.2 than at pH 7.7. The endonuclease formed single-strand breaks making 5'-phosphoryl and 3'-hydroxyl termini, and deoxythymidine was present at the 5' termini with a frequency of about 50% in both optimum conditions. Bovine pancreatic DNase I antibody and G-action inhibited the enzyme activity. Thus this endonuclease is classified as a DNase I.
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PMID:Purification and properties of a neutral endodeoxyribonuclease from rat small intestinal mucosa. 707 91

An endonuclease active upon single-stranded DNA has been extensively purified from Ustilago maydis. The native form of the enzyme appears to be a single polypeptide chain of 55,000 daltons. The enzyme does not require addition of divalent cations for activity, but is stimulated severalfold by the transition metals Co2+, Mn2+, and Zn2+. It is strongly inhibited by ethylenediaminetetraacetic acid and 2-mercaptoethanol. The products of reaction are mononucleotides and small oligonucleotides bearing a 5'-phosphomonoester group. Superhelical DNA is cleaved by the enzyme to an open circular form, then cut to form a linear molecule. Relaxed closed circular duplex DNA is resistant to cleavage. Heteroduplex relaxed circular DNA molecules containing nonhomology or damage in only one strand were constructed and tested as substrates for the enzyme. The rate of enzymatic cleavage was found to be high relative to a control when molecules containing depurinated, deaminated, or radiation damaged sites were tested. Molecules containing a single mismatched base pair were not cleaved any faster than a completely base-paired control, unless reactions were carried out under conditions likely to introduce additional structural distortions in the DNA. Linear duplex DNA exposed to the enzyme is progressively shortened by digestion from both 5' and 3' termini.
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PMID:Studies on nuclease alpha from Ustilago maydis. 722 71

We have purified a novel endonuclease from Escherichia coli that recognizes deoxyinosine, a deamination product of deoxyadenosine in DNA. This activity, which we named deoxyinosine 3' endonuclease, is different from the known hypoxanthine DNA N-glycosylases which have been partially characterized in E. coli and other organisms. The enzyme was purified 24,800-fold to apparent homogeneity. SDS- and activity PAGE analyses indicate that the enzyme has an apparent molecular mass of 25 kDa. Deoxyinosine 3' endonuclease recognized deoxyinosine in both single- and double-stranded DNA but exhibited a 4-fold preference for double stranded over single-stranded DNA. In addition to deoxyinosine, the enzyme recognized urea residues and AP sites. Deoxyinosine 3' endonuclease has an obligatory requirement for Mg2+, but other cations such as Co2+ and Mn2+ could partially replace Mg2+. The optimal pH for deoxyinosine 3' endonuclease was around 7.5. In contrast to most of the known repair enzymes, deoxyinosine 3' endonuclease makes an incision at the second phosphodiester bond 3' to a deoxyinosine or AP site, leaving behind the intact lesion on the nicked DNA. Therefore, deoxyinosine 3' endonuclease recognizes, but does not remove, the lesion from the DNA molecule. The biological significance of this novel activity is discussed with reference to other repair activities in E. coli.
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PMID:Purification and characterization of a novel deoxyinosine-specific enzyme, deoxyinosine 3' endonuclease, from Escherichia coli. 820 31


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