Genetic Incorporation of Selenotyrosine Significantly Improves Enzymatic Activity of Agrobacterium radiobacter Phosphotriesterase

Tyrosine plays important roles in many enzymes. To facilitate enzyme design, mechanistic studies and minimize structural perturbation in the active site, here we report the genetic incorporation of a novel unnatural amino acid selenotyrosine (SeHF), which has single‐atom replacement in comparison to...

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Bibliographic Details
Published in:Chembiochem : a European journal of chemical biology 2021-08, Vol.22 (15), p.2535-2539
Main Authors: An, Xiaojing, Chen, Chao, Wang, Tianyuan, Huang, Aiping, Zhang, Dawei, Han, Ming‐Jie, Wang, Jiangyun
Format: Article
Language:English
Subjects:
Agrobacterium radiobacter
Agrobacterium tumefaciens - enzymology
Agrobacterium tumefaciens - genetics
Amino acids
Catalytic Domain
Enzymatic activity
Enzyme activity
Enzymes
Hydrogen-Ion Concentration
Kinetics
Molecular dynamics
Molecular Dynamics Simulation
Mutation
Perturbation
pH effects
Phosphoric Triester Hydrolases - chemistry
Phosphoric Triester Hydrolases - genetics
Phosphoric Triester Hydrolases - metabolism
Phosphotriesterase
phosphotriesterases
selenotyrosine
Tyrosine
Tyrosine - analogs & derivatives
Tyrosine - chemistry
Tyrosine - metabolism
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Summary:Tyrosine plays important roles in many enzymes. To facilitate enzyme design, mechanistic studies and minimize structural perturbation in the active site, here we report the genetic incorporation of a novel unnatural amino acid selenotyrosine (SeHF), which has single‐atom replacement in comparison to tyrosine. The arPTE‐(Agrobacterium radiobacter Phosphotriesterase) Tyr309SeHF mutant exhibits a significant 12‐fold increase in kcat and 3.2‐fold enhancement in kcat/KM at pH 7.0. Molecular dynamics simulations show that the SeHF309 mutation results in a conformational switch which opens up the product release pocket and increases the product release rate, thereby elevating the overall enzyme activity. Significant improvement of the catalytic efficiency at neutral pH by single unnatural amino acid (UAA) mutation broadens the application of this enzyme, and provides valuable insights to the mechanism. Our method represents a new approach for designing enzymes with enhanced activity. Tyrosine mimic, amino acid SeHF, which has low pKa and strong nucleophilicity under physiological conditions, could significantly improve the enzymatic activity of Agrobacterium radiobacter phosphotriesterase. This approach could be used in metalloenzyme design in the future.
ISSN:1439-4227
1439-7633
1439-7633
DOI:10.1002/cbic.202000460