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On the active site of mononuclear B1 metallo β-lactamases: a computational study

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Abstract

Metallo-β-lactamases (MβLs) are Zn(II)-based bacterial enzymes that hydrolyze β-lactam antibiotics, hampering their beneficial effects. In the most relevant subclass (B1), X-ray crystallography studies on the enzyme from Bacillus Cereus point to either two zinc ions in two metal sites (the so-called ‘3H’ and ‘DCH’ sites) or a single Zn(II) ion in the 3H site, where the ion is coordinated by Asp120, Cys221 and His263 residues. However, spectroscopic studies on the B1 enzyme from B. Cereus in the mono-zinc form suggested the presence of the Zn(II) ion also in the DCH site, where it is bound to an aspartate, a cysteine, a histidine and a water molecule. A structural model of this enzyme in its DCH mononuclear form, so far lacking, is therefore required for inhibitor design and mechanistic studies. By using force field based and mixed quantum–classical (QM/MM) molecular dynamics (MD) simulations of the protein in aqueous solution we constructed such structural model. The geometry and the H-bond network at the catalytic site of this model, in the free form and in complex with two common β-lactam drugs, is compared with experimental and theoretical findings of CphA and the recently solved crystal structure of new B2 MβL from Serratia fonticola (Sfh-I). These are MβLs from the B2 subclass, which features an experimentally well established mono-zinc form, in which the Zn(II) is located in the DCH site. From our simulations the εεδ and δεδ protomers emerge as possible DCH mono-zinc reactive species, giving a novel contribution to the discussion on the MβL reactivity and to the drug design process.

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Notes

  1. A short MD calculation (400 ps) has been run removing the postion restraints on the water molecules after the equilibration, however Wat-H conserves its propensity to exchange its position with other water molecules from the bulk. Then the structure equilibrated with postion restraints was used in the subsequent QM/MM MD.

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Acknowledgments

This work has been supported in part by the FIRB project Contract RBLA032ZM7, by the EC contract SPINE II n° 031220 and by grants from HHMI (Howard Hughes Medical Instituteand ANPCyT (Agencia Nacional de Promoción Científica y Tecnológica) to AJV. AJV is a fellow of the John Simon Guggenheim Foundation and an HHMI International Research Scholar. Access to the computational resources supplied by CASPUR and by CINECA is gratefully acknowledged.

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Author notes

  1. Jacopo Sgrignani

    Present address: CNR-IOM-DEMOCRITOS National Simulation Center at SISSA, via Bonomea 265, Trieste, Italy

Authors and Affiliations

  1. CERM and Department of Chemistry “Ugo Schiff”, University of Florence, Via Luigi Sacconi 6, 50019, Sesto Fiorentino, Italy

    Jacopo Sgrignani & Roberta Pierattelli

  2. CNR-IOM-DEMOCRITOS National Simulation Center at SISSA, via Bonomea 265, Trieste, Italy

    Alessandra Magistrato

  3. Laboratory for Biomolecular Modeling, Institute of Bioengineering, School of Life Sciences, Ecole Polytechnique Fédérale de Lausanne—EPFL, 1015, Lausanne, Switzerland

    Matteo Dal Peraro

  4. Instituto de Biología Molecular y Celular de Rosario (IBR); Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), Facultad de Ciencias Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario (UNR), Suipacha 531, S2002LRK, Rosario, Argentina

    Alejandro J. Vila

  5. Computational Biophysics, German Research School for Simulation Sciences, 52425, Jülich, Germany

    Paolo Carloni

  6. Institute for Advanced Simulation, Forschungszentrum Jülich, 52425, Jülich, Germany

    Paolo Carloni

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  1. Jacopo Sgrignani
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Correspondence to Alessandra Magistrato or Roberta Pierattelli.

Additional information

Paolo Carloni: Joint venture of RWTH Aachen University and Forschungszentrum Jülich, Germany.

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Sgrignani, J., Magistrato, A., Dal Peraro, M. et al. On the active site of mononuclear B1 metallo β-lactamases: a computational study. J Comput Aided Mol Des 26, 425–435 (2012). https://doi.org/10.1007/s10822-012-9571-0

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