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| In vitro cleavage of hepatitis B virus C mRNA by 10-23 DNA enzyme |
| Wei Hou, Jian-Er Wo, Min-Wei Li and Ke-Zhou Liu |
Hangzhou, China
Author Affiliations: Institute of Infectious Diseases, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou 310003, China (Hou W, Wo JE, Li MW and Liu KZ)
Corresponding Author: Jian-Er Wo, PhD, Institute of Infectious Diseases, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou 310003, China (Tel: 86-571-87236579; Fax: 86-571-87068731; Email: wojianer@zju.edu.cn) |
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Abstract BACKGROUND: 10-23 DNA enzyme is one kind of deoxyribozymes for RNA cleavage. The inhibition effects of 10-23 DNA enzyme on the expression of the HBV C gene in HepG2.2.15 cells were demonstrated previously. The aim of this study was to further explore the cleavage activities of 10-23 DNA enzyme targeting at HBV C gene mRNA in vitro.
METHODS: 10-23 DNA enzyme named Drz-HBV-C-9 specific to HBV C gene ORF A1816UG was designed and synthesized. HBV C gene mRNA was obtained by the in vitro transcription method. Cleavage activities of Drz-HBV-C-9 were observed in vitro. Values of kinetic parameters including Km，Kcat and Kcat/Km were calculated accordingly.
RESULTS: Under the certain cleavage conditions, Drz-HBV-C-9 could efficiently cleave target mRNA at specific sites in vitro. Cleavage products of 109nt plus 191nt were obtained. The kinetic parameters, Km，Kcat and Kcat/Km for Drz-HBV-C-9, were 1.4×10-9 mol, 1.6 min-1 and 1.1×109 mol-1•min-1, respectively.
CONCLUSIONS: 10-23 DNA enzyme targeting at HBV C gene mRNA possesses specific cleavage activities in vitro. This would be a potent antiviral strategy with respect to HBV gene therapy.
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2005, Vol. 4 
