Effects of Divalent Mg2+ and Monovalent Na+ on Cleavage Reactions by Multiribozyme System in vitro
- VernacularTitle:Mg2+和Na+对多核酶系统体外切割反应的影响
- Author:
Shengli TIAN
;
Shuo ZHENG
;
Shide LIU
;
Dongping XU
;
Takao OHNUMA
- Publication Type:Journal Article
- Keywords:
muhifibozyme system;
divalent Mg2+;
monovalent Na+;
RNA cleavage;
cleavage reaction
- From:
Chinese Journal of Biochemistry and Molecular Biology
2008;24(7):612-618
- CountryChina
- Language:Chinese
-
Abstract:
To better understand the cleavage efficiency of muhiribozyme system on its RNA substrate in the presence and absence of divalent magnesium and monovalent sodium ions.we constructed pGEM-Coat'A,pGEM-Coat'A196Rz plasmids and pGEM-MDRl target plasmid.They were applied to transcribe RNAs with SP6/T7 transcription kit.Cleavage reactions were carried out in cell-free system and reaction products were analyzed by electrophoresis on 6% denaturing polyacrylamide gels in TBS buffer.The gels were dried and exposed to X-ray films for autoradiography.The Image J software was employed to analyze the dried gels.The results indicated that the cleavage efficiency of the muhiribozyme was dependent on the concentration of divalent Mg2+.The cleavage products increased with the concentrations of divalent Mg2+ and were Mg2+ concentration and time dependent.No cleavage product was obtained in the presence of lower than 200 mmol/L Na+ alone.On the contrary,monovalent Na+ inhibited the Mg2+ -induced cleavage reaction in Na+ and Mg2+ coexistance.The cleavage rate was significantly lower than that observed with divalent Mg2+ alone.These results suggested that divalent Mg2+ was required for muhiribozyme on substrate cleavage reaction in the physical condition,whereas monovalent Na+ was not.
