The single chain antibody scFv2F3 can be converted into selenium-containing Se-scFv2F3 by chemical mutation of the Ser residues. With antibody fragment 1NQB as a template, the catalytic domain of scFv2F3 was built by using homology modeling and molecular dynamics（MD） simulations. On the basis of the 3D model, we discussed the importance of Ser52 as the chemical modification site and redesigned the protein groups nearby Ser52 via intro- ducing a catalytic triad. The following 10 ns MD results show that the designed Ser52-Trp29-Gln72 catalytic triad is stable enough and high close to the local structural features of native glutathione peroxidases（GPX）. Our results may be useful for creating a new abzyme with higher catalytic efficiency and stability.
The cytochrome P450 mutant CYP2C9.13（L90P） shows a greatly impaired catalytic activity compared with the wild-type. We constructed the mutants by substitution at residue 90 of CYP2C9, expressed in COS-7 cells, assayed their thermal stability and catalysis activity and analyzed the mutants via molecular dynamic（MD） simulation and flexible docking. Mutant L90E exhibits a significantly lower catalytic activity than the wild-type for the hydroxylation of diclofenac, lornoxicam and luciferin and its molecular dynamics simulation results indicate that the size of the entrance of substrate access was reduced significantly. An increase or minor decrease of catalytic activity was observed for mutants L90Q, L90W, L90R, L90I and L90G, and the sizes of the entrances of substrate access and the active site cavities had a little change in those mutants. The thermal stability and the potential energy of the MD simulation of these mutants showed a similar tendency as the catalysis assays did. Flexible docking results show the fluctuation of interaction energy is due to the change of electrostatic potential distribution. All the above facts show that the changes in the catalysis activity of the mutants caused by the substitution at residue 90 are due to the changes in the size of entrance, the shape and size of active site cavity, electrostatic potential distribution and thermal stability. The residue 90 of CYP2C9 has an important effect on the enzyme catalytic activity.
WANG YingZHOU Yi-hanGUO Ying-jieXU Xue-lianSI Da-yongZHOU HuiLI Ze-sheng
The density functional theory at the B3LYP/6-311G（d, p） level was applied to exploring the inhibition mechanism of cholinesterases by carbamate. The results indicate that the inhibition reactions with or without the catalytic effect of the catalytic triad in eholinesterases underwent a two-step addition-elimination mechanism, which is in good agreement with the proposed mechanism. The solvent has a strong effect on the inhibition reactions and the reaction with the catalytic triad in the solvent phase is close to the real reaction under biological condition.
