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Cristina Romero-López, Alicia Barroso-delJesus, Elena Puerta-Fernández, Alfredo Berzal-Herranz

Interfering with hepatitis C virus IRES activity using RNA molecules identified by a novel in vitro selection method

Keywords: anti-hepatitis C virus internal ribosome entry site (HCV IRES) RNAs, catalytic RNAs, hepatitis C virus internal ribosome entry site (HCV IRES), inhibitor RNAs, in vitro selection, RNA aptamers

Hepatitis C virus (HCV) infection is one of the world's major health problems, and the identification of efficient HCV inhibitors is a major goal. Here we report the isolation of efficient anti-HCV internal ribosome entry site (IRES) RNA molecules identified by a new in vitro selection method. The newly developed procedure consists of two sequential steps that use distinct criteria for selection: selection for binding and selection for cleaving. The selection protocol was applied to a population of more than 10¹⁵ variants of an anti-hepatitis C virus ribozyme covalently linked to an aptamer motif. The ribozyme was directed against positions 357 to 369 of the HCV IRES, and the cleavage substrate was a 691-nucleotide-long RNA fragment that comprises the entire HCV IRES domain. After six selection cycles, seven groups of RNA variants were identified. A representative of each group was tested for its capacity to inhibit IRES activity using in vitro translation assays. All selected RNAs promoted significant inhibition, some by as much as 95%.

Biological Chemistry, Walter de Gruyter

Print ISSN: 1431-6730
Volume: 386, 02/2005
Pages: 183 - 190

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