Cleavage of influenza RNA by using a human PUF-based artificial RNA-binding protein–staphylococcal nuclease hybrid

Tomoaki Mori, Kento Nakamura, Keisuke Masaoka, Yusuke Fujita, Ryosuke Morisada, Koichi Mori, Takamasa Tobimatsu, Takashi Sera

Research output: Contribution to journalArticle

Abstract

Various viruses infect animals and humans and cause a variety of diseases, including cancer. However, effective methodologies to prevent virus infection have not yet been established. Therefore, development of technologies to inactivate viruses is highly desired. We have already demonstrated that cleavage of a DNA virus genome was effective to prevent its replication. Here, we expanded this methodology to RNA viruses. In the present study, we used staphylococcal nuclease (SNase) instead of the PIN domain (PilT N-terminus) of human SMG6 as an RNA-cleavage domain and fused the SNase to a human Pumilio/fem-3 binding factor (PUF)-based artificial RNA-binding protein to construct an artificial RNA restriction enzyme with enhanced RNA-cleavage rates for influenzavirus. The resulting SNase-fusion nuclease cleaved influenza RNA at rates 120-fold greater than the corresponding PIN-fusion nuclease. The cleaving ability of the PIN-fusion nuclease was not improved even though the linker moiety between the PUF and RNA-cleavage domain was changed. Gel shift assays revealed that the RNA-binding properties of the PUF derivative used was not as good as wild type PUF. Improvement of the binding properties or the design method will allow the SNase-fusion nuclease to cleave an RNA target in mammalian animal cells and/or organisms.

Original languageEnglish
Pages (from-to)736-740
Number of pages5
JournalBiochemical and Biophysical Research Communications
Volume479
Issue number4
DOIs
Publication statusPublished - Oct 28 2016

Fingerprint

RNA Cleavage
Micrococcal Nuclease
Human Influenza
RNA
Viruses
Fusion reactions
DNA Viruses
RNA-Binding Proteins
RNA Viruses
Virus Diseases
Animals
Gels
Genome
Technology
Enzymes
Assays
Neoplasms
Genes
Cells
Derivatives

Keywords

  • Artificial RNA restriction enzyme
  • RNA cleavage
  • RNA virus

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Cleavage of influenza RNA by using a human PUF-based artificial RNA-binding protein–staphylococcal nuclease hybrid. / Mori, Tomoaki; Nakamura, Kento; Masaoka, Keisuke; Fujita, Yusuke; Morisada, Ryosuke; Mori, Koichi; Tobimatsu, Takamasa; Sera, Takashi.

In: Biochemical and Biophysical Research Communications, Vol. 479, No. 4, 28.10.2016, p. 736-740.

Research output: Contribution to journalArticle

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