Structure of RNA-dependent RNA polymerase from 2019-nCoV, a major antiviral drug target

Yan Gao; Liming Yan; Yucen Huang; Fengjiang Liu; Yao Zhao; Lin Cao; Tao Wang; Qianqian Sun; Zhenhua Ming; Lianqi Zhang; Ji Ge; Litao Zheng; Ying Zhang; Haofeng Wang; Yan Zhu; Chen Zhu; Tianyu Hu; Tian Hua; Bing Zhang; Xiuna Yang; Jun Li; Haitao Yang; Zhijie Liu; Wenqing Xu; Luke W. Guddat; Quan Wang; Zhiyong Lou; Zihe Rao

All dose-fractioned images were motion-corrected and dose-weighted by MotionCorr2 40 software (22) and their contrast transfer functions were estimated by cryoSPARC patch CTF estimation. For Dataset-1, a total of 2,334,248 particles were auto-picked using blob picker and extracted with a box size of 300 pixels in cryoSPARC (20). The following 2D, 3D classifications and refinements were all performed in cryoSPARC. 918,133 particles were selected after two rounds of 2D classification. 100,000 particles were used to do Ab-Initio reconstruction in five classes, and then these five classes were used as 3D volume templates for heterogeneous refinement with all selected particles, with 110,176 particles converged into one class. Next, this particle set was used to perform homogeneous refinement, yielding a resolution of 3.1 Ã…. After local refinement, the final resolution reached 2.9 Ã…. For Dataset-2, the image processing was conducted using a similar pipeline. 753,481 particles were auto-picked initially, and 145,388 5 particles were selected after final heterogeneous refinement. The resolution reached 2.99 Ã… after non-uniform refinement and 2.95 Ã… after local refinement with a mask.