Software for protein engineering

Current version: 4.0.0
Released: MAR/17/2008

Description

The methodology of computational site-directed mutagenesis and the in silico testing of mutant properties in program TRITON consists of following steps:

  • Modelling protein mutant structures
    The 3D structures of mutants are generated by the MODELLER program. This program uses the method of comparative protein modling by satisfaction of spatial restrains to model structures of homology proteins based on the 3D structure of template protein (typicaly X-ray structure from structural database) and the amino acid sequence of modeled protein.
    Scheme of protein mutants modeling
  • Modelling reaction pathway
    The modeled structures of mutant proteins are used for calculation of reaction pathway. Only the active site residues, substrate, co-substrate, and co-factors are included in the calculation. In order to mimic the situation in the enzyme, positions of all backbone atoms are fixed.
    The reaction pathway is calculated by the semiempirical quantum chemistry program MOPAC. The subroutine DRIVER is used for reaction pathway mapping. The procedure produces data which can be used to visualise the relationships between the energy and the reaction coordinate, and to estimate the activation barrier and the thermodynamics of the reaction. Changes in partial charges on individual atoms are also monitored during the calculation.
    Output data can be analyded in graphical interface of TRITON. The energies and partial charges on atoms are obtained from output files. The activation barriers of the reaction, changes in partial atomic charges and electrostatic interactions of active site residues with substrate are then calculated. This data can be used for assessment of enzyme activity.
    Scheme of MOPAC/DRIVER computational steps
  • Modelling ligand-protein affinity
    Modelled structures of mutant proteins are used for docking of ligand to protein. External program AutoDock is used for generation of a set of ligand-protein complexes. Graphical interface of TRITON is used to add hydrogen atoms, charges and solvatation parameters to structure. Subsequently grid box and AutoGrid/Autodock parameters are specified.
    After finishing AutoDock calculation, protein-ligand complexes can be displayed. Also electrostatic interaction of ligand with individual residues can be evaluated which can describe influence of this residues to afinity. Visualisation of affinity maps cam help to find areas with highest affinity toward individual elements.
    Scheme of AutoDock computational steps

 
 

Last update: March 17, 2008