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Software for protein engineering
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Current version: 4.0.0
Released: MAR/17/2008
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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.
- 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.
- 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.
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