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Engineering the Pseudomonas aeruginosa II lectin: designing mutants with changed affinity and specificity

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Abstract

This article focuses on designing mutations of the PA-IIL lectin from Pseudomonas aeruginosa that lead to change in specificity. Following the previous results revealing the importance of the amino acid triad 22–23–24 (so-called specificity-binding loop), saturation in silico mutagenesis was performed, with the intent of finding mutations that increase the lectin’s affinity and modify its specificity. For that purpose, a combination of docking, molecular dynamics and binding free energy calculation was used. The combination of methods revealed mutations that changed the performance of the wild-type lectin and its mutants to their preferred partners. The mutation at position 22 resulted in 85 % in inactivation of the binding site, and the mutation at 23 did not have strong effects thanks to the side chain being pointed away from the binding site. Molecular dynamics simulations followed by binding free energy calculation were performed on mutants with promising results from docking, and also at those where the amino acid at position 24 was replaced for bulkier or longer polar chain. The key mutants were also prepared in vitro and their binding properties determined by isothermal titration calorimetry. Combination of the used methods proved to be able to predict changes in the lectin performance and helped in explaining the data observed experimentally.

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Acknowledgments

This work was supported by the Czech Science Foundation (Contract No. 13-25401S), the Ministry of Education, Youth and Sports of the Czech Republic (Contract No. LH13055), The European Community’s Seventh Framework Programme under European Regional Development Fund (Contract No. CZ.1.05/1.1.00/02.0068) and under the “Capacities” specific programme (Contract No. 286154—SYLICA). The access to MetaCentrum supercomputing facilities provided under the research intent LM2010005 is highly appreciated.

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Authors and Affiliations

  1. CEITEC – Central European Institute of Technology, Masaryk University, Kamenice 5, 625 00, Brno, Czech Republic

    Zdeněk Kříž, Jan Adam, Michaela Wimmerová & Jaroslav Koča

  2. National Centre for Biomolecular Research, Faculty of Science, Masaryk University, Brno, Czech Republic

    Zdeněk Kříž, Jan Adam, Jana Mrázková, Petros Zotos, Thomais Chatzipavlou, Michaela Wimmerová & Jaroslav Koča

  3. Division of Pharmaceutical Chemistry, School of Pharmacy, National and Capodistrian University of Athens, Athens, Greece

    Petros Zotos & Thomais Chatzipavlou

  4. Department of Biochemistry, Faculty of Science, Masaryk University, Brno, Czech Republic

    Jana Mrázková & Michaela Wimmerová

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  1. Zdeněk Kříž
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  2. Jan Adam
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  3. Jana Mrázková
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Correspondence to Michaela Wimmerová or Jaroslav Koča.

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Zdeněk Kříž, Jan Adam and Jana Mrázková have equally contributed to this work.

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Kříž, Z., Adam, J., Mrázková, J. et al. Engineering the Pseudomonas aeruginosa II lectin: designing mutants with changed affinity and specificity. J Comput Aided Mol Des 28, 951–960 (2014). https://doi.org/10.1007/s10822-014-9774-7

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  • DOI: https://doi.org/10.1007/s10822-014-9774-7

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