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Identification and characterization of an unusual metallo-β-lactamase from Serratia proteamaculans

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Abstract

Metallo-β-lactamases (MBLs) are a family of metalloenzymes that are capable of hydrolyzing β-lactam antibiotics and are an important means by which bacterial pathogens use to inactivate antibiotics. A database search of the available amino acid sequences from Serratia proteamaculans indicates the presence of an unusual MBL. A full length amino acid sequence alignment indicates overall homology to B3-type MBLs, but also suggests considerable variations in the active site, notably among residues that are relevant to metal ion binding. Steady-state kinetic measurements further indicate functional differences and identify two relevant pK a values for catalysis (3.8 for the enzyme–substrate complex and 7.8 for the free enzyme) and a preference for penams with modest reactivity towards some cephalosporins. An analysis of the metal ion content indicates the presence of only one zinc ion per active site in the resting enzyme. In contrast, kinetic data suggest that the enzyme may operate as a binuclear enzyme, and it is thus proposed that a catalytically active di-Zn2+ center is formed only once the substrate is present.

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Abbreviations

ICP-MS:

Inductively coupled plasma mass spectrometry

MBL:

Metallo-β-lactamases

PAP:

Purple acid phosphatase

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Acknowledgments

This work was supported by a National Health and Medical Research Council of Australia Project Grant and an SFI President of Ireland Young Researcher Award. G.S. acknowledges the award of an Australian Research Council Future Fellowship (FT120100694).

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Authors and Affiliations

  1. School of Chemistry and Molecular Biosciences, The University of Queensland, St. Lucia, QLD, 4072, Australia

    Peter Vella, Eleanor W. W. Leung, Fernanda Ely, Ross P. McGeary & Gerhard Schenk

  2. Department of Chemistry, National University of Ireland, Maynooth, Maynooth, Co. Kildare, Ireland

    Manfredi Miraula, Emer Phelan & Nataša Mitić

  3. Research School of Chemistry, The Australian National University, Canberra, ACT, 0200, Australia

    David L. Ollis

  4. School of Pharmacy, The University of Queensland, St. Lucia, QLD, 4072, Australia

    Ross P. McGeary

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  1. Peter Vella
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Correspondence to Gerhard Schenk or Nataša Mitić.

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Vella, P., Miraula, M., Phelan, E. et al. Identification and characterization of an unusual metallo-β-lactamase from Serratia proteamaculans . J Biol Inorg Chem 18, 855–863 (2013). https://doi.org/10.1007/s00775-013-1035-z

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