title: Unique 5'-P recognition and basis for dG:dGTP misincorporation of ASFV DNA polymerase X.
abstract: African swine fever virus (ASFV) can cause highly lethal disease in pigs and is becoming a global threat. ASFV DNA Polymerase X (AsfvPolX) is the most distinctive DNA polymerase identified to date; it lacks two DNA-binding domains (the thumb domain and 8-KD domain) conserved in the homologous proteins. AsfvPolX catalyzes the gap-filling reaction during the DNA repair process of the ASFV virus genome; it is highly error prone and plays an important role during the strategic mutagenesis of the viral genome. The structural basis underlying the natural substrate binding and the most frequent dG:dGTP misincorporation of AsfvPolX remain poorly understood. Here, we report eight AsfvPolX complex structures; our structures demonstrate that AsfvPolX has one unique 5'-phosphate (5'-P) binding pocket, which can favor the productive catalytic complex assembly and enhance the dGTP misincorporation efficiency. In combination with mutagenesis and in vitro catalytic assays, our study also reveals the functional roles of the platform His115-Arg127 and the hydrophobic residues Val120 and Leu123 in dG:dGTP misincorporation and can provide information for rational drug design to help combat ASFV in the future.
authors.affiliation: State Key Laboratory of Genetic Engineering, Collaborative Innovation Center of Genetics and Development, Department of Physiology and Biophysics, School of Life Sciences, Fudan University, Shanghai, China.
authors.forename: Yiqing
authors.initials: Y
authors.lastname: Chen
authors.affiliation: State Key Laboratory of Genetic Engineering, Collaborative Innovation Center of Genetics and Development, Department of Physiology and Biophysics, School of Life Sciences, Fudan University, Shanghai, China.
authors.forename: Jing
authors.initials: J
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Genetic Engineering, Collaborative Innovation Center of Genetics and Development, Department of Physiology and Biophysics, School of Life Sciences, Fudan University, Shanghai, China.|State Key Laboratory of Genetic Engineering, Collaborative Innovation Center of Genetics and Development, Department of Biochemistry, Institute of Plant Biology, School of Life Sciences, Fudan University, Shanghai, China.
authors.forename: Hehua
authors.initials: H
authors.lastname: Liu
authors.affiliation: State Key Laboratory of Genetic Engineering, Collaborative Innovation Center of Genetics and Development, Department of Physiology and Biophysics, School of Life Sciences, Fudan University, Shanghai, China.
authors.forename: Yanqing
authors.initials: Y
authors.lastname: Gao
authors.affiliation: State Key Laboratory of Genetic Engineering, Collaborative Innovation Center of Genetics and Development, Department of Biochemistry, Institute of Plant Biology, School of Life Sciences, Fudan University, Shanghai, China.
authors.forename: Xuhang
authors.initials: X
authors.lastname: Li
authors.affiliation: State Key Laboratory of Genetic Engineering, Collaborative Innovation Center of Genetics and Development, Department of Biochemistry, Institute of Plant Biology, School of Life Sciences, Fudan University, Shanghai, China.
authors.forename: Lina
authors.initials: L
authors.lastname: Zheng
authors.affiliation: State Key Laboratory of Genetic Engineering, Collaborative Innovation Center of Genetics and Development, Department of Biochemistry, Institute of Plant Biology, School of Life Sciences, Fudan University, Shanghai, China.
authors.forename: Ruixue
authors.initials: R
authors.lastname: Cui
authors.affiliation: State Key Laboratory of Genetic Engineering, Collaborative Innovation Center of Genetics and Development, Department of Biochemistry, Institute of Plant Biology, School of Life Sciences, Fudan University, Shanghai, China.
authors.forename: Qingqing
authors.initials: Q
authors.lastname: Yao
authors.affiliation: State Key Laboratory of Genetic Engineering, Collaborative Innovation Center of Genetics and Development, Department of Biochemistry, Institute of Plant Biology, School of Life Sciences, Fudan University, Shanghai, China.
authors.forename: Liang
authors.initials: L
authors.lastname: Rong
authors.affiliation: State Key Laboratory of Genetic Engineering, Collaborative Innovation Center of Genetics and Development, Department of Physiology and Biophysics, School of Life Sciences, Fudan University, Shanghai, China.
authors.forename: Jixi
authors.initials: J
authors.lastname: Li
authors.affiliation: Department of Chemistry, Georgia State University, Atlanta, Georgia, United States of America.|College of Life Sciences, Sichuan University, Chengdu, China.
authors.forename: Zhen
authors.initials: Z
authors.lastname: Huang
authors.affiliation: State Key Laboratory of Genetic Engineering, Collaborative Innovation Center of Genetics and Development, Department of Biochemistry, Institute of Plant Biology, School of Life Sciences, Fudan University, Shanghai, China.
authors.forename: Jinbiao
authors.initials: J
authors.lastname: Ma
authors.affiliation: State Key Laboratory of Genetic Engineering, Collaborative Innovation Center of Genetics and Development, Department of Physiology and Biophysics, School of Life Sciences, Fudan University, Shanghai, China.
authors.forename: Jianhua
authors.initials: J
authors.lastname: Gan
mesh_terms: D000358:African Swine Fever Virus / Q000201:enzymology*; D000596:Amino Acids / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites; D018360:Crystallography, X-Ray; D004279:DNA, Viral / Q000737:chemistry / Q000378:metabolism; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism*; D003848:Deoxyguanine Nucleotides / Q000378:metabolism*; D007700:Kinetics; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D017433:Protein Structure, Secondary; D040681:Structural Homology, Protein
chemical_list: D000596:Amino Acids; D004279:DNA, Viral; D003848:Deoxyguanine Nucleotides; D050505:Mutant Proteins; C029603:deoxyguanosine triphosphate; C109534:DNA polymerase X; D004259:DNA-Directed DNA Polymerase