Degradation of DNA into 5'-Monodeoxyribonucleotides in the Presence of Mn^<2+> Ions

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概要

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• DNA is known to be aggregated by metal ions including Mn2+ ions, but analysis of the aggregation process from a chemical viewpoint, which means identification of the product yielded during the process, has not been performed yet. On examination of the kinds of degraded materials that were in the supernatant obtained on centrifugation of a DNA mixture aggregated under conditions of 10 mM Mn2+ ions ([Mn]/[P] = 46.3) at 70 °C for 1 h, the degradation products were found to be dAMP, dCMP, dGMP, and TMP. These dNMPs were purified by HPLC on TSKgel ODS-80Ts and identified by LC-TOF/MS. The degradation activity was lost on pretreatment of the DNA with a phenol–chloroform mixture, and the activity was recovered by pretreatment with a mixture of DMSO and a buffer containing surfactants. Mn2+, Co2+, Ni2+, Cu2+, Zn2+, and Cd2+, as transition element metal ions, were effective as to the degradation into dNMP. Mg2+, Ca2+, Sr2+, and Ba2+, as alkali earth element metal ions, were not effective as to the degradation. Monovalent anions such as Cl−, CH3OO−, and NO3− were found to increase the degradation rate. Sixty μg of the 120 μg of the starting DNA in 450 μl was degraded into dNMP on reaction for 1 h in the presence of 100 mM NaCl and 10 mM Mn2+ ions. In this process, aggregation did not occur, and thus was not considered to be necessary for degradation. The degradation was found not to occur at pH 7.0, and to be very sensitive to pH. The OH− ion should have a critical role in cleavage of the phosphodiester linkages in this case. The dNMP obtained in the degradation process was found to be only 5′-NMP, based on the H1NMR spectra. This prosess should prove to be a new process for the production of 5′-dNMP in addtion to the exonuclease.

• 2007-11-23

著者

• Maeda Hidekatsu

  Division of Bioengineering, Graduate School of Engineering, Soka University

• Ikeguchi Masamichi

  Division of Bioengineering, Graduate School of Engineering, Soka University

• UEKI Shouji

  Faculty of Pharmaceutical Sciences, Kagawa Campus, Tokushima Bunri University

• Ueki Shouji

  Faculty Of Pharmaceutical Sciences Kagawa Campus Tokushima Bunri University

• Arata Toshiaki

  Division Of Biological Sciences Graduate School Of Science Osaka University

• Wada Shinya

  Division Of Bioengineering Graduate School Of Engineering Soka University

• Ikeguchi Masamichi

  Division Of Bioengineering Graduate School Of Engineering Soka University

• Minoura Norihiko

  School Of Bionics Tokyo University Of Technology

• Maeda Hidekatsu

  Div. Of Bioinformatics Graduate School Of Engineering Soka Univ.

• Maeda Hidekatsu

  Division Of Bioengineering Graduate School Of Engineering Soka University

関連論文

• Degradation of DNA into 5′-Monodeoxyribonucleotides in the Presence of Mn2+ Ions
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