Previously, we reported that our designed artificial DNA-cleaving enzyme composed of an artificial DNA-binding protein designed to specifically bind to a target DNA sequence and a DNA-cleaving domain prevented DNA replication of human papilloma virus (HPV) in mammalian cells by sequence-specifically cleaving its target DNA sequence in an HPV genome. One of the advantages of our methodology for inactivating DNA viruses is that it can be applied to any DNA virus by adjusting its DNA binding domain of our artificial DNA-cleaving enzyme for viruses of interest. Here, we expanded the methodology for inactivating DNA viruses to RNA viruses. In the present study, to inhibit RNA replication of influenza virus, we developed artificial RNA-cleaving enzymes composed of a sequence-specific RNA-binding domain based on a human Pumilio/fem-3 binding factor and an RNA-cleaving domain via four flexible peptide linkers different in length. We determined the best peptide for RNA cleavage and demonstrated that the resulting artificial enzyme site-selectively and efficiently cleaved its target RNA sequence in an influenza viral genome in vitro. Next, we examined whether the artificial RNA-cleaving enzymes efficiently cleaved their target RNA sequence and inhibited RNA replication of influenza virus in mammalian cells. We will report the results in detail.

Development of novel artificial RNA-cleaving enzymes for inactivating RNA viruses

ACS Spring Meeting 2021

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