• 제목/요약/키워드: chemical nuclease

검색결과 16건 처리시간 0.027초

$^{32}$P-Postlabelling 방법의 응용 : Azo색소 및 Flavonoid화합물에 의해 유도되는 DNA Adduct의 겸출에 관한 연구 (Application of the $^{32}$P-Postlabelling Technique : A Study on Detection of DNA Adduct Induced by Azo Dyes rind Flavonoid Compounds)

  • 김재현;박창원;박정식;홍연탁;김효정;이주한;이헌수;이동권
    • Biomolecules & Therapeutics
    • /
    • 제1권1호
    • /
    • pp.58-64
    • /
    • 1993
  • DNA addicts induced by putative chemical related to carcinogenesis were detected and determined by $^{32}$P-Postlabelling assay after exposure of 4 compounds comprising two auto dyes (amaranth, new coccine) and two flavonoid compounds (rutin, quercetin) to ICR mouse. DNA was isolated from mouse liver and digested enzymatically to deoxyribonucleoside 3'-monophosphate. The postincubation of DNA digests with nuclease Pl before $^{32}$P-labelling enhanced the technique's sensitivity. Nuclease Pl cleaves deoxyribonucleoside 3'-mono-phosphates of normal nucleotides to deoxyrihonucleosides which do not serve as substrates for polynucleotide kinase, while most of addicts were found to be totally or partially resistant to the 3'-dephosphorylating action of nuclease Pl. The adducted deoxyribonucleoside 3'-monophosphate was converted to 5'-$^{32}$P-labelled deoxynucleoside 3',5'-bisphosphate by T4 polynucleotide kinase. The nucleotides were separated by anion-exchange thin layer chromatography(TLC) on polyethyleneimine cellulose by 4-dimensional or 2-dimensional TLC then detected by autoradiography. The results show that DNA addicts were detected in liver DNA of ICR mouse after administration of amaranth and quercetin by 2-dimensional and/or 4-dimensional TLC.

  • PDF

Human ChlR1 Stimulates Endonuclease Activity of hFen1 Independently of ATPase Activity

  • Kim, Do-Hyung;Kim, Jeong-Hoon;Park, Byoung Chul;Lee, Do Hee;Cho, Sayeon;Park, Sung Goo
    • Bulletin of the Korean Chemical Society
    • /
    • 제35권10호
    • /
    • pp.3005-3008
    • /
    • 2014
  • Human ChlR1 protein (hChlR1), a member of the cohesion establishment factor family, plays an important role in the segregation of sister chromatids for maintenance of genome integrity. We previously reported that hChlR1 interacts with hFen1 and stimulates its nuclease activity on the flap-structured DNA substrate covered with RPA. To elucidate the relationship between hChlR1 and Okazaki fragment processing, the effect of hChlR1 on in vitro nuclease activities of hFen1 and hDna2 was examined. Independent of ATPase activity, hChlR1 stimulated endonuclease activity of hFen1 but not that of hDna2. Our findings suggest that the acceleration of Okazaki fragment processing near cohesions may aid in reducing the size of the replication machinery, thereby facilitating its entry through the cohesin ring.

Detection of Cleavage Sites on 5S rRNA by Methidiumpropyl-EDTA-Iron(II)

  • Kim, Sang-Bumn;Cho, Bong-Rae;Lee, Young-Hoon;Park, In-Won
    • BMB Reports
    • /
    • 제29권2호
    • /
    • pp.133-136
    • /
    • 1996
  • The affinity cleavage reagent Methidiumpropyl-EDTA-Iron(II) is applied to the structural analysis of 5S rRNA. Analysis of cleavage sites induced by MPE-Fe(II) on 5S rRNA shows that MPE intercalates easily between the unstable base pairs or into the bulges, thereby it strongly cuts the nucleosides nearby. The stable helical stems A, B, D and E as well as loop d are weakly cut. Most of the single-stranded loops are not cleaved. Based on the cleavage pattern of the 5S rRNA by MPE-Fe(II) and RNase V1, we suggest that MPE-Fe(II) may be used as a potential chemical probe in searching for the unstable helical regions of RNA, and for the sequences that appear to be involved in folding and distorting 5S rRNA.

  • PDF

Synthesis and Configuration Analysis of Diastereomers of 5'-O-(2'-Deoxycytidyl)-3'-O-Thymidyl Phosphorothioate

  • 문병조;정현주;김상국;김남희
    • Bulletin of the Korean Chemical Society
    • /
    • 제17권1호
    • /
    • pp.24-28
    • /
    • 1996
  • A procedure is described for the synthesis of the title compound via phosphotriester intermediates. The preparation of $R_p$ and $S_p$ diastereomeric dinucleotide of d[Cp(S)T] was performed by the condensation of the protected deoxycytidine, the protected thymidine, 2,5-dichlorophenylphosphorodichloridothioate and 1-hydroxybenzotriazole in THF. Their designation of configuration at phosphorus as $R_p$ and $S_p$ follows from anaylsis of ${31}^P$ NMR spectroscopy and reverse-phase HPLC and the stereospecificity in the hydrolysis catalyzed by Nuclease S1 and snake venom phosphodiesterase. Diastereomerically pure $R_p$ and $S_p$ d[Cp(S)T] were utilized to synthesize oligonucleotides containing the XhoI recognition sequence with a phosphorothioate group at the cleavage site.