In silico saturation mutagenesis and docking screening for the analysis of protein-ligand interaction: the Endothelial Protein C Receptor case study

Chiappori Federica, D'Ursi Pasqualina, Merelli Ivan, Milanesi Luciano, Rovida, Rovida Ermanna

Description

The design of mutants in protein functional regions, such as the ligand binding sites, is a powerful approach to recognize the determinants of specific protein activities in cellular pathways. For an exhaustive analysis of selected positions of protein structure large scale mutagenesis techniques are often employed, with laborious and time consuming experimental set-up. 'In silico' mutagenesis and screening simulation represents a valid alternative to laboratory methods to drive the 'in vivo' testing toward more focused objectives. Results We present here a high performance computational procedure for large-scale mutant modelling and subsequent evaluation of the effect on ligand binding affinity. The mutagenesis was performed with a 'saturation' approach, where all 20 natural amino acids were tested in positions involved in ligand binding sites. Each modelled mutant was subjected to molecular docking simulation and stability evaluation. The simulated protein-ligand complexes were screened for their impairment of binding ability based on change of calculated Ki compared to the wild-type. An example of application to the Endothelial Protein C Receptor residues involved in lipid binding is reported. Conclusion The computational pipeline presented in this work is a useful tool for the design of structurally stable mutants with altered affinity for ligand binding, considerably reducing the number of mutants to be experimentally tested. The saturation mutagenesis procedure does not require previous knowledge of functional role of the residues involved and allows extensive exploration of all possible substitutions and their pairwise combinations. Mutants are screened by docking simulation and stability evaluation followed by a rationally driven selection of those presenting the required characteristics. The method can be employed in molecular recognition studies and as a preliminary approach to select models for experimental testing.

Informations

• Publié par: biomed
• Publié le: 01 janvier 2009
• Nombre de lectures: 7
• Langue: English

Extrait

BMC

Bioinformatics

BioMedCentral

Open Access Research In silico saturation mutagenesis and docking screening for the analysis of proteinligand interaction: the Endothelial Protein C Receptor case study 1,2 1,2 1 1 Federica Chiappori , Pasqualina D’Ursi , Ivan Merelli , Luciano Milanesi 1 and Ermanna Rovida*

1 Address: Institute for Biomedical Technologies–National Research Council (ITBCNR), via Fratelli Cervi 93, 20090 Segrate (MI), Italy and 2 Consorzio Interuniversitario Lombardo per l’Elaborazione Automatica (CILEA), via Raffaello Sanzio 4, 20090 Segrate (MI), Italy Email: Federica Chiappori  [e-mail]; Pasqualina D’Ursi  [e-mail]; Ivan Merelli  [e-mail]; Luciano Milanesi  [e-mail]; Ermanna Rovida*  [e-mail] *Corresponding author

fromBioinformatics Methods for Biomedical Complex Systems Applications (NETTAB2008) Varenna, Italy 19–21 May 2008

Published: 15 October 2009 BMC Bioinformatics2009,10(Suppl 12):S3

doi: 10.1186/1471210510S12S3

This article is available from: http://www.biomedcentral.com/14712105/10/S12/S3 ©2009 Chiappori et al; licensee BioMed Central Ltd. This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract Background:The design of mutants in protein functional regions, such as the ligand binding sites, is a powerful approach to recognize the determinants of specific protein activities in cellular pathways. For an exhaustive analysis of selected positions of protein structure large scale mutagenesis techniques are often employed, with laborious and time consuming experimental set up.‘In silico’mutagenesis and screening simulation represents a valid alternative to laboratory methods to drive the‘in vivo’testing toward more focused objectives. Results:We present here a high performance computational procedure for largescale mutant modelling and subsequent evaluation of the effect on ligand binding affinity. The mutagenesis was performed with a‘saturation’approach, where all 20 natural amino acids were tested in positions involved in ligand binding sites. Each modelled mutant was subjected to molecular docking simulation and stability evaluation. The simulated proteinligand complexes were screened for their impairment of binding ability based on change of calculated Ki compared to the wildtype. An example of application to the Endothelial Protein C Receptor residues involved in lipid binding is reported. Conclusion:The computational pipeline presented in this work is a useful tool for the design of structurally stable mutants with altered affinity for ligand binding, considerably reducing the number of mutants to be experimentally tested. The saturation mutagenesis procedure does not require previous knowledge of functional role of the residues involved and allows extensive exploration of all possible substitutions and their pairwise combinations. Mutants are screened by

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