Crystal structures of PigF, an O-methyltransferase involved in the prodigiosin synthetic pathway, reveal an induced-fit substrate-recognition mechanism
Shenshen Qiu,
Dongqing Xu,
Mengxue Xu,
Huan Zhou,
Ning Sun,
Li Zhang,
Mengmeng Zhao,
Jianhua He,
Tingting Ran,
Bo Sun,
Weiwu Wang
Affiliations
Shenshen Qiu
Department of Microbiology, College of Life Sciences, Nanjing Agricultural University, Nanjing 210095, People's Republic of China
Dongqing Xu
Department of Microbiology, College of Life Sciences, Nanjing Agricultural University, Nanjing 210095, People's Republic of China
Mengxue Xu
Department of Microbiology, College of Life Sciences, Nanjing Agricultural University, Nanjing 210095, People's Republic of China
Huan Zhou
Shanghai Synchrotron Radiation Facility, Shanghai Advanced Research Institute, Chinese Academy of Sciences, Shanghai 201204, People's Republic of China
Ning Sun
Department of Microbiology, College of Life Sciences, Nanjing Agricultural University, Nanjing 210095, People's Republic of China
Li Zhang
Department of Microbiology, College of Life Sciences, Nanjing Agricultural University, Nanjing 210095, People's Republic of China
Mengmeng Zhao
Department of Microbiology, College of Life Sciences, Nanjing Agricultural University, Nanjing 210095, People's Republic of China
Jianhua He
Shanghai Synchrotron Radiation Facility, Shanghai Advanced Research Institute, Chinese Academy of Sciences, Shanghai 201204, People's Republic of China
Tingting Ran
Department of Microbiology, College of Life Sciences, Nanjing Agricultural University, Nanjing 210095, People's Republic of China
Bo Sun
Shanghai Synchrotron Radiation Facility, Shanghai Advanced Research Institute, Chinese Academy of Sciences, Shanghai 201204, People's Republic of China
Weiwu Wang
Department of Microbiology, College of Life Sciences, Nanjing Agricultural University, Nanjing 210095, People's Republic of China
Prodigiosin, a red linear tripyrrole pigment, is a typical secondary metabolite with numerous biological functions, such as anticancer, antibacterial and immunosuppressant activities, and is synthesized through a bifurcated biosynthesis pathway from 4-methoxy-2,2′-bipyrrole-5-carbaldehyde (MBC) and 2-methyl-3-n-amylpyrrole (MAP). The last step in the biosynthetic pathway of MBC is catalysed by PigF, which transfers a methyl group to 4-hydroxy-2,20-bipyrrole-5-carbaldehyde (HBC) to form the final product MBC. However, the catalytic mechanism of PigF is still elusive. In this study, crystal structures of apo PigF and S-adenosylhomocysteine (SAH)-bound PigF were determined. PigF forms a homodimer and each monomer consists of two domains: a C-terminal catalytic domain and an N-terminal dimerization domain. Apo PigF adopts an open conformation, while the structure of the complex with the product SAH adopts a closed conformation. The binding of SAH induces dramatic conformational changes of PigF, suggesting an induced-fit substrate-binding mechanism. Further structural comparison suggests that this induced-fit substrate-recognition mechanism may generally exist in O-methyltransferases. Docking and mutation studies identified three key residues (His98, His247 and Asp248) that are crucial for enzyme activity. The essential function of His247 and Asp248 and structure analysis suggests that both residues are involved in activation of the HBC substrate of PigF. The invariance of Asp248 in PigF further confirmed its essential role. The invariance and essential role of His98 in PigF suggests that it is involved in correctly positioning the substrate. This study provides new insight into the catalytic mechanism of PigF, reveals an induced-fit substrate-recognition model for PigF and broadens the understanding of O-methyltransferases.
