科研之友
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摘要
Acetylcholinesterase (AChE) has been widely used for the detection of organophosphate and carbamate pesticides, due to its high sensitivity and low limit of detection to the presence of pesticides. The homology modeled recombinant Bombyx mori Acetylcholinesterase II (rBm-AChE II) and docking results with multiple pesticides inferred that Y398, located at the bottleneck of the active site gorge, might be important for enzyme sensitivity. Thus, three mutants (Y398G, Y398F, Y398W) were constructed and exhibited different enzyme activities and sensitivities. The results showed that Y398W possessed a remarkably increased enzyme activity, while Y398F had a significant reduction. The Y398F has an approximately 2-fold lower IC50 for some pesticides than the wild type enzyme, indicating a higher sensitivity. With the detailed investigation of the conformations of computer simulation, we propose that for the positively charged and small substrate ATChI, a larger side chain at position 398 improves the fixation of the substrate in an appropriate conformation for catalysis. For bulky substrates such as pesticides, the diffusion in the active site gorge may be related to the enlargement of the bottleneck by having proper orientations more easily. In addition, a more hydrophobic side chain at the bottleneck seemed to be beneficial for ligand diffusion.
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