Research Articles

C2c2 is a single-component programmable RNA-guided RNA-targeting CRISPR effector

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Science  05 Aug 2016:
Vol. 353, Issue 6299, aaf5573
DOI: 10.1126/science.aaf5573

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  • Are Guide DNAs functioning at physiological temperature range possible for Argonaute or CRISPR/Cas9 system?
    • Qiuyun Liu, Superviser/Writting/Professor, State Key Laboratory of Biocontrol, MOE Key Laboratory of Aquatic Product Safety, Sun Yat-sen University, China
    • Other Contributors:
      • Yulin Wan, Writting/Student, School of Life Sciences, Sun Yat-sen University, Guangzhou 510275, China
      • Yunfan Shi, Writting/Student, School of Life Sciences, Sun Yat-sen University, Guangzhou 510275, China
      • Feng Wang, Writting/Professor, Faculty of Science, Engineering and Technology, Swinburne University of Technology, Australia
      • Xiaoyi Hu, Writting/Student, Department of Chemical and Biomolecular Engineering, New York University Tandon School of Engineering, USA

    Guide RNAs are used in CRISPR/Cas9 systems (1), and Guide DNAs could function in some prokaryotic Argonaute systems at high temperatures (2-3), which are well above physiological temperature ranges. RNA possesses extra hydroxyl groups that are nucleophilic, and can also form secondary chemical bonding with cations. Therefore, a guide DNA functioning at physiological temperature range requires the introductions of nucleophilic amino acid residues such as serine and threonine into the relevant proteins, in order to enhance nucleophilic catalysis as well as to attract cations. Due to the electron-withdrawing feature of the gamma-methyl group in threonine compared with serine according to NMR data, the nucleophilic catalysis with the hydroxyl oxygen of serine may be more potent than that of threonine. Leucine to valine replacements could also be beneficial as the carbonyl oxygen of valine possesses affinities to cations (4) to provide a similar milieu as offered by guider RNAs. The potential toxicity associated with generated gene variants during protein engineering may be addressed by the incorporation of acidic amino acids, alanine, glycine, etc. (5).

    Yulin Wan, 1 Yunfan Shi, 1 Feng Wang, 2 Xiaoyi Hu, 3 Qiuyun Liu, 1*
    1. State Key Laboratory of Biocontrol, MOE Key Laboratory of Aquatic Product Safety, The Key Laboratory of Gene Engineering of Ministry of Education, School of Life Sciences, Sun Yat-sen University, Guangzhou, 510275, China.
    2. Molecular Mo...

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    Competing Interests: None declared.

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