Acta Phys. -Chim. Sin. ›› 2013, Vol. 29 ›› Issue (02): 439-448.doi: 10.3866/PKU.WHXB201211272

• BIOPHYSICAL CHEMISTRY • Previous Articles    

Rational Design of Affinity Ligand for the Oriented Immobilization of Trypsin

BAI Shu, ZHOU Rong, LIU Fu-Feng   

  1. Department of Biochemical Engineering, School of Chemical Engineering and Technology, Tianjin University, Tianjin 300072, P. R. China; Key Laboratory of Systems Bioengineering, Ministry of Education, Tianjin University, Tianjin 300072, P. R. China
  • Received:2012-09-20 Revised:2012-11-19 Published:2013-01-14
  • Supported by:

    The project was supported by the National Natural Science Foundation of China (20906068), Natural Science Foundation of Tianjin from Tianjin Municipal Science and Technology Commission, China (10JCYBJC04500).


Based on the three-dimensional (3D) structure of trypsin, 2-nitrophenyl-β-D-glucopyranoside was selected from the ZINC database to be a candidate affinity ligand for trypsin. The affinity between trypsin and the ligand was analyzed. It is found that the interactions between the ligand and the protein are dominated by van der Waals interactions and hydrogen bonding. Molecular dynamics (MD) simulations were used to verify the affinity between the ligand and trypsin; the simulations indicate that the complex remains stable, and the distance between the ligand and the target protein changes only a little. It is found that one water molecule acts as a bridge between the ligand and the protein pocket via hydrogen bonding. Finally, the ligand was coupled to Sepharose CL-6B gel, and was used to immobilize trypsin in an oriented fashion. It is found that the enzyme activity and specific activity of the oriented immobilized trypsin are 340.8 U·g-1 and 300.3 U·mg-1, respectively. These values are 10 and 5 times that of the free enzyme. The results of this work indicate that the combination of docking and MD simulations are promising for the rational design of ligands for oriented immobilization.

Key words: Enzyme catalysis, Affinity ligand, Molecular docking, Molecular dynamics simulation, Oriented immobilization