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Updated: Feb 2, 2026

In Vitro Directed Evolution of a Restriction Endonuclease with More Stringent Specificity
Published on: March 25, 2020
Characterization of an endonuclease in rice stripe tenuivirus Pc1 in vitro
Shuling Zhao1, Guodong Xu1, Guanghui He1
1College of Bioscience and Biotechnology, Yangzhou University, Yangzhou, 225009, People's Republic of China; Joint International Research Laboratory of Agriculture & Agri-Product Safety, Yangzhou University, People's Republic of China.
Abstract:
Rice stripe tenuivirus (RSV) initiates its mRNA transcription by using the cap-snatching mechanism during which an endonuclease activity is required for the cleavage of host mRNA. In this study, we aim to characterize the endonuclease in RSV. Sequence alignment revealed the presence of a cap-snatching endonuclease domain in RSV Pc1. Expression and in vitro enzymatic activity assay demonstrated that this domain indeed had a manganese-dependent endonuclease activity. The enzyme could efficiently degrade ssRNA with preference for unstructured ssRNA, but not DNA. Mutations in the endonuclease domain allowed the identification of four key residues (D547, D567, E585 and K604). The endonuclease of RSV was similar but not identical to other known viral endonucleases, suggesting that RSV endonuclease may have some distinct catalytic characteristics.
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