Triple mutated antibody scFv2F3 with high GPx activity: insights from MD, docking, MDFE, and MM-PBSA simulation

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• 作者：Quan Luo (1)
  Chunqiu Zhang (1)
  Lu Miao (1)
  Dongmei Zhang (1)
  Yushi Bai (1)
  Chunxi Hou (1)
  Junqiu Liu (1)
  Fei Yan (2)
  Ying Mu (2)
  Guimin Luo (2)
  
• 关键词：Abzyme ; GPx mimic ; Docking ; MD simulation ; MDFE simulation ; MM ; PBSA
• 刊名：Amino Acids
• 出版年：2013
• 出版时间：March 2013
• 年：2013
• 卷：44
• 期：3
• 页码：1009-1019
• 全文大小：823KB
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• 作者单位：Quan Luo (1)
  Chunqiu Zhang (1)
  Lu Miao (1)
  Dongmei Zhang (1)
  Yushi Bai (1)
  Chunxi Hou (1)
  Junqiu Liu (1)
  Fei Yan (2)
  Ying Mu (2)
  Guimin Luo (2)
  
  1. State Key Laboratory of Supramolecular Structure and Materials, Institute of Theoretical Chemistry, Jilin University, Changchun, 130012, People’s Republic of China
  2. Key Laboratory for Molecular Enzymology and Engineering of the Ministry of Education, Jilin University, Changchun, People’s Republic of China
  
• ISSN：1438-2199

文摘

By combining computational design and site-directed mutagenesis, we have engineered a new catalytic ability into the antibody scFv2F3 by installing a catalytic triad (Trp29–Sec52–Gln72). The resulting abzyme, Se-scFv2F3, exhibits a high glutathione peroxidase (GPx) activity, approaching the native enzyme activity. Activity assays and a systematic computational study were performed to investigate the effect of successive replacement of residues at positions 29, 52, and 72. The results revealed that an active site Ser52/Sec substitution is critical for the GPx activity of Se-scFv2F3. In addition, Phe29/Trp–Val72/Gln mutations enhance the reaction rate via functional cooperation with Sec52. Molecular dynamics simulations showed that the designed catalytic triad is very stable and the conformational flexibility caused by Tyr101 occurs mainly in the loop of complementarity determining region 3. The docking studies illustrated the importance of this loop that favors the conformational shift of Tyr54, Asn55, and Gly56 to stabilize substrate binding. Molecular dynamics free energy and molecular mechanics-Poisson Boltzmann surface area calculations estimated the pK a shifts of the catalytic residue and the binding free energies of docked complexes, suggesting that dipole–dipole interactions among Trp29–Sec52–Gln72 lead to the change of free energy that promotes the residual catalytic activity and the substrate-binding capacity. The calculated results agree well with the experimental data, which should help to clarify why Se-scFv2F3 exhibits high catalytic efficiency.