An Argonaute from Thermus parvatiensis exhibits endonuclease activity mediated by 5′chemically modified DNA guides  

作　　者：Yingying Sun Xiang Guo Hui Lu Liuqing Chen Fei Huang Qian Liu Yan Feng 

机构地区：[1]State Key Laboratory of Microbial Metabolism,School of Life Sciences and Biotechnology,Shanghai Jiao Tong University,Shanghai 200240,China [2]Shenzhen Institutes of Advanced Technology,Chinese Academy of Sciences,Shenzhen 518055,China

出　　处：《Acta Biochimica et Biophysica Sinica》2022年第5期686-695,共10页生物化学与生物物理学报（英文版）

基　　金：supported by the grants from the Natural Science Foundation of China(No.31770078);the Ministry of Science and Technology(No.2020YFA0907700).

摘　　要：Prokaryotic Argonaute(pAgo)nucleases with precise DNA cleavage activity show great potential for gene manipulation.Extensive biochemical studies have revealed that recognition of guides with different 5′groups by Ago is important for biocatalysis.Here,we identified an Ago from the thermophilic Thermus parvatiensis(TpsAgo)and analyzed the regulatory effect of 5′-modified guides on TpsAgo cleavage activity.Recombinant TpsAgo cleaves single-stranded DNA and RNA targets at 65–90°C,which is mediated by a 5′hydroxyl or phosphate DNA guide.Notably,TpsAgo can utilize various 5′-modified DNA guides for catalysis,including 5′-NH2C6,5′-Biotin,5′-FAM and 5′-SHC6 guides.Moreover,TpsAgo performs programmable cleavage of double-stranded DNA at temperatures over 80°C and strongly tolerates NaCl concentrations up to 3.2 M.These results provide insight into the catalytic performance of Agos by guide regulation,which may facilitate their biotechnological applications.

关 键 词：ARGONAUTE Thermus parvatiensis ENDONUCLEASE 5′-modified guide 

分 类 号：O64[理学—物理化学]