• BMB Reports
• /
• v.31 no.4
• /
• pp.405-408
• /
• 1998

Endonuclease-sensitive sites detected by T4 endonuclease V or UvrABC nuclease treatments were compared in the dihydrofolate reductase gene of UV-irradiated Chinese hamster ovary B-11 cells. The number of endonuclease-sensitive sites detected by T4 endonuclease V treatment followed by NaOH denaturation was twice that of formamide denaturation. Repeated treatment of damaged genomic DNA with T4 endonuclease V resulted in no further increase in the number of endonuclease-sensitive sites detected. The numbers of endonuclease-sensitive sites detected by UvrABC nuclease using each denaturation condition were similar. Sequential treatment with the two endonucleases using formamide denaturation resulted in twice the number of endonuclease-sensitive sites detected by treatment of each nuclease alone. Due to a lack of AP endonuclease activity these results suggest the presence of T4 endonuclease V-sensitive sites which could be complemented by alkaline gel separation or by UvrABC nuclease treatment.

• Microbiology and Biotechnology Letters
• /
• v.15 no.5
• /
• pp.352-355
• /
• 1987

The isolation and characterization of type II restriction endonuclease from Clostridium thermocellum ATCC 27405 were described. This enzyme (Cth I endonuclease) is an isoschizomer of Bcl I endonuclease recognizing 5'-TGATCA-3'. Cth I endonuclease requires MG$^{2+}$ ion for its activity and is maximally active at PH 1.5 to 10.5 in the Presence of 0 to 10mM NaCl. Cth I endonuclease is heat stable and has an optimum temperature of 6$0^{\circ}C$. The activity of Cth I enzyme is sensitive to dam methylation.

• Korean Journal of Microbiology
• /
• v.24 no.4
• /
• pp.406-410
• /
• 1986

The isolation and charateriation of a type II restriction endonuclease from Xanthomonas citri IFO 3835 were described. This enzyme (Xci I endonuclease) is an isoschizomer of Sal I endonuclease recognizing 5'-GTCGAC-3' and cleaving at the site indicated by the arrow. Unlike Sal I endonuclease, Xci I endonuclease requries a NaCl concentration of 50mM for its maximum activity.

• BMB Reports
• /
• v.31 no.1
• /
• pp.106-110
• /
• 1998

We have employed a DNA-native-polyacrylamide gel electrophoresis (DNA-native-PAGE) assay system to characterize the enzyme activity of the endonuclease secreted by human B lymphoblastic IM9 cells. Experimental results clearly demonstrated that the endonuclease activity of IM9 cell culture medium is distinct from that of DNase I in the DNA-native-PAGE assay system. Immunoprecipitation analysis using anti-DNase I antibody showed that the secreted endonuclease is not recognized by the antibody. The secreted endonuclease was estimated using supercoiled plasmid DNA as a substrate. The pH optimum required for the catalytic activity was determined to be in the range of pH 6.6-7.4. No significant difference in the endonuclease secretion was observed by stimulation of the IM9 cells with interferon-${\gamma}$ or interleukin-$1{\beta}$.

• YAKHAK HOEJI
• /
• v.40 no.2
• /
• pp.193-201
• /
• 1996

In order to study the role of C-terminal aromatic region of T4 endonuclease V in the interaction with substrate DNA, NMR and Fluorescence spectrum were recorded. Analysis of flu orescence emission spectra showed that C-terminal region of T4 endonuclease V is in or very near the binding site. In the HSQC spectrum of $^{15}N$-Tyr-labeled T4 endonuclease V*DNA complex, the broadening of a peak was observed. It is presumed that this peak corresponds to one among three tyrosine residues which belong to the WYKYY segment of C-terminal region of T4 endonuclease V. Interactions of peptide fragments consisting of C-terminal residues of T4 endonuclease V with DNAs(TT-, T^T-DNA) were investigated by NMR and Fluorescence experiment. The results suggest that two peptide fragments themselves bind to DNAs and their binding pattern is not an intercalation mode.

• BMB Reports
• /
• v.30 no.1
• /
• pp.66-72
• /
• 1997

A mammalian DNA repair enzyme, UV endonuclease III which also functions as a ribosomal protein S3 (rpS3), was purified from mouse cells and characterized. UV endonuclease III was previously cloned and known to yield a peptide of 32 kDa upon expression in E. coli [Kim et al., (1995) J. Bioi. Chem. 270, 13620-13629]. However, biochemically purified UV endonuclease III, which has a sedimentation coefficient of 3.25, appears to have an additional peptide of 28 kDa. It appears that two bands were derived from one complex, judging from the comparison of the nuclease activity on the native and SDS-gel electrophoreses. UV endonuclease III becomes non-specific upon purification and this phenomenon is more significant in the case of pure fractions of the enzyme. Non-specific activity was not influenced by pH or any salt conditions.

• Korean Journal of Microbiology
• /
• v.26 no.1
• /
• pp.1-5
• /
• 1988

The esolation and characterization of a new type II restriction endounclease, BamI, from Bacellus macerans ATCC 8244 were described. BmaI endonuclease was partially purified by procedures of ammonium sulfate fractionation, DEAE-cellulose and phosphocellulose chromatographies. This enzume recognized one site on pBR322 DNA, two sites on Bluescribe DNA, three sites on $\lambda$DNA and no site on SV 40 DNA. The same cleavage patterns for vareius DNAs as PvuI indicated that BamI is an isoschisomer of PvuI whose recognition sequence is 5'-CGATCG-3'. The optimal pH for the BmaI endonuclease activity was about 7.0 and optimal NaCl concentration was about 100mM. Manganese ion could partially replace magnesium as a cofactor, but calcium could not at all.

• Journal of Microbiology and Biotechnology
• /
• v.33 no.6
• /
• pp.788-796
• /
• 2023

Canine parvovirus type 2 (CPV-2) has high morbidity and mortality rates in canines. Nonstructural protein 1 (NS1) of CPV-2 has endonuclease activity, initiates viral DNA replication, and is highly conserved. Thus, it is a promising target for antiviral inhibitor development. We overexpressed a 41.9 kDa active recombinant endonuclease in Escherichia coli and designed a nicking assay using carboxyfluorescein and quencher-linked ssDNA as substrates. The optimal temperature and pH of the endonuclease were 37℃ and pH 7, respectively. Curcumin, bisdemethoxycurcumin, demethoxycurcumin, linoleic acid, tannic acid, and α-tocopherol inhibited CPV-2 NS1 endonuclease with IC₅₀ values of 0.29 to 8.03 µM. The extracted turmeric, yerba mate, and sesame cake suppressed CPV-2 NS1 endonuclease with IC₅₀ values of 1.48, 7.09, and 52.67 ㎍/ml, respectively. The binding affinity between curcumin, the strongest inhibitor, and CPV-2 NS1 endonuclease by molecular docking was -6.4 kcal/mol. Curcumin inhibited CPV-2 NS1 endonuclease via numerous hydrophobic interactions and two hydrogen bonds with Lys97 and Pro111 in the allosteric site. These results suggest that adding curcuminoids, linoleic acid, tannic acid, α-tocopherol, extracted turmeric, sesame cake, and yerba to the diet could prevent CPV-2 infection.

• Journal of the Korean Society of Tobacco Science
• /
• v.10 no.1
• /
• pp.37-47
• /
• 1988

Endonuclease inhibitor was isolated from the chloroplast of the leaves of tobacco, Burley 21 and NC 83, The inhibitory effect on the activities of endonucleases including BamH1 was lost by the addition of trypsin or heat treatment However, the treatment of $\alpha$-amylase was unconcerned in the inhibitory effect The endonuclease inhibitor was found to be a monomeric protein that plays a vital role for protection of tobacco chloroplast DNA from endonuclease action.

• Journal of Plant Biology
• /
• v.33 no.4
• /
• pp.293-301
• /
• 1990

We have purified and characterized a deoxyribonuclease from zygotes of the eukaryotic green alga, Chlamydomonas reinhardtii and investigated effects of the polyamines, putrescine, spermidine and spermine on the purifed endonuclease-catalyzed cleavage of plasmid DNA. The enzyme has a molecular weight of about 37 kDa as measured by gel filtration and SDS-polyacrylamide gel electrophoresis. There is no requirement for a divalent cation. The activity is sensitive to ionic strength, as NaCl and KCl result in inhibition. The cleavage of plasmid DNA by the purified endonuclease was effectively inhibited by polyamines. The enzyme activity was inhibited more effectively by spermine than by spermidine. The inhibition by putrescine was lower than the other two polyamines.