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Evolution of a new enzyme for carbon disulphide conversion by an acidothermophilic archaeon

Nature volume 478, pages 412–416 (2011)Cite this article

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Abstract

Extremophilic organisms require specialized enzymes for their exotic metabolisms. Acid-loving thermophilic Archaea that live in the mudpots of volcanic solfataras obtain their energy from reduced sulphur compounds such as hydrogen sulphide (H2S) and carbon disulphide (CS2)1,2. The oxidation of these compounds into sulphuric acid creates the extremely acidic environment that characterizes solfataras. The hyperthermophilic Acidianus strain A1-3, which was isolated from the fumarolic, ancient sauna building at the Solfatara volcano (Naples, Italy), was shown to rapidly convert CS2 into H2S and carbon dioxide (CO2), but nothing has been known about the modes of action and the evolution of the enzyme(s) involved. Here we describe the structure, the proposed mechanism and evolution of a CS2 hydrolase from Acidianus A1-3. The enzyme monomer displays a typical β-carbonic anhydrase fold and active site, yet CO2 is not one of its substrates. Owing to large carboxy- and amino-terminal arms, an unusual hexadecameric catenane oligomer has evolved. This structure results in the blocking of the entrance to the active site that is found in canonical β-carbonic anhydrases and the formation of a single 15-Å-long, highly hydrophobic tunnel that functions as a specificity filter. The tunnel determines the enzyme’s substrate specificity for CS2, which is hydrophobic. The transposon sequences that surround the gene encoding this CS2 hydrolase point to horizontal gene transfer as a mechanism for its acquisition during evolution. Our results show how the ancient β-carbonic anhydrase, which is central to global carbon metabolism, was transformed by divergent evolution into a crucial enzyme in CS2 metabolism.

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Figure 1: Neighbour-joining phylogenetic tree showing the evolutionary relationship between carbonic anhydrases (CAs) and the newly discovered CS 2 hydrolases.
Figure 2: X-ray crystal structure of the Acidianus A1-3 CS 2 hydrolase.
Figure 3: The CS 2 hydrolase active site and the entrance to the active site.
Figure 4: Effect of mutations in the access tunnel on activity.

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Accession codes

Primary accessions

GenBank/EMBL/DDBJ

Protein Data Bank

Data deposits

The sequence for the CS2 hydrolase from Acidianus A1-3 has been deposited in GenBank under the accession number HM805096. Atomic coordinates and structure factor amplitudes have been deposited in the Protein Data Bank under accession numbers 3TEO and 3TEN.

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Acknowledgements

J. Eygensteyn is acknowledged for ICP-MS analysis. We thank H. Harhangi for advice on cloning, S. Zimmermann for assistance with cloning, A. Meinhart for discussions and support for cloning and crystallography, and D. Ringe and J. Reinstein for discussions. We thank the Dortmund-Heidelberg data collection team, especially W. Blankenfeldt, and the staff of beam lines X10SA and X12SA at the Swiss Light Source of the PSI in Villigen for their help and facilities. We also thank A. Rufer for help with the analytical ultracentrifuge and I. Vetter for support with the crystallographic software. The work was funded by an STW grant (STW_6353) to M.J.S. and M.H.Z. and by the Max-Planck Society.

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Author notes

  1. Marjan J. Smeulders and Thomas R. M. Barends: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Microbiology, Radboud University Nijmegen, Heyendaalseweg 135, 6525 AJ, Nijmegen, The Netherlands

    Marjan J. Smeulders, Arjan Pol, Marcel H. Zandvoort, Ahmad F. Khadem, John Hermans, Lina Russ, Mike S. M. Jetten & Huub J. M. Op den Camp

  2. Department of Biomolecular Mechanisms, Max-Planck Institute for Medical Research, Jahnstrasse 29, D-69120 Heidelberg, Germany

    Thomas R. M. Barends, Anna Scherer, Anikó Udvarhelyi, Robert L. Shoeman & Ilme Schlichting

  3. Paul Scherrer Institut, CH-5232 Villigen PSI, Switzerland

    Andreas Menzel

  4. Department of Laboratory Medicine, Nijmegen Centre for Mitochondrial Disorders, Nijmegen Proteomics Facility, Radboud University Nijmegen Medical Centre, Geert Grooteplein 10PO Box 9101, 6500 HB Nijmegen, The Netherlands

    Hans J. C. T. Wessels & Lambert P. van den Heuvel

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Contributions

The research was conceived by A.P., M.S.M.J. and H.J.M.O. M.H.Z. performed the sampling, enrichment and isolation. M.J.S., M.H.Z. and A.P. performed the physiological experiments. M.J.S., J.H., A.F.K., L.P.V. and H.J.C.T.W. performed the purification and protein and gene sequencing. A.F.K. and H.J.M.O. performed the MALDI-TOF MS. H.J.M.O. and M.J.S. performed the alignments and phylogenetic analyses. M.J.S. and L.R. performed the site-directed mutant studies, including the activity measurements. A.S. and T.R.M.B. grew the crystals; T.R.M.B. determined the crystal structures and suggested the amino acid residues to be mutated; I.S., A.U. and A.M. performed the SAXS experiments and analyses; and R.L.S. performed the analytical ultracentrifugation experiments. M.J.S., T.R.M.B., I.S., M.S.M.J. and H.J.M.O. wrote the manuscript. All authors discussed the results and commented on the manuscript.

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Correspondence to Mike S. M. Jetten.

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Smeulders, M., Barends, T., Pol, A. et al. Evolution of a new enzyme for carbon disulphide conversion by an acidothermophilic archaeon. Nature 478, 412–416 (2011). https://doi.org/10.1038/nature10464

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