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Structural basis for synthesis of inflammatory mediators by human leukotriene C4 synthase

Nature volume 448, pages 613–616 (2007)Cite this article

Abstract

Cysteinyl leukotrienes are key mediators in inflammation and have an important role in acute and chronic inflammatory diseases of the cardiovascular and respiratory systems, in particular bronchial asthma. In the biosynthesis of cysteinyl leukotrienes, conversion of arachidonic acid forms the unstable epoxide leukotriene A4 (LTA4). This intermediate is conjugated with glutathione (GSH) to produce leukotriene C4 (LTC4) in a reaction catalysed by LTC4 synthase1: this reaction is the key step in cysteinyl leukotriene formation. Here we present the crystal structure of the human LTC4 synthase in its apo and GSH-complexed forms to 2.00 and 2.15 Å resolution, respectively. The structure reveals a homotrimer, where each monomer is composed of four transmembrane segments. The structure of the enzyme in complex with substrate reveals that the active site enforces a horseshoe-shaped conformation on GSH, and effectively positions the thiol group for activation by a nearby arginine at the membrane–enzyme interface. In addition, the structure provides a model for how the ω-end of the lipophilic co-substrate is pinned at one end of a hydrophobic cleft, providing a molecular ‘ruler’ to align the reactive epoxide at the thiol of glutathione. This provides new structural insights into the mechanism of LTC4 formation, and also suggests that the observed binding and activation of GSH might be common for a family of homologous proteins important for inflammatory and detoxification responses.

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Figure 1: LTC 4 synthase overall structure.
Figure 2: Substrate–protein and lipid–protein interactions.
Figure 3: Glutathione binding.
Figure 4: The hydrophobic binding crevice and proposed schematic mechanism of substrate binding and catalysis.

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Acknowledgements

This work was supported by the Swedish Research Council, the Swedish Cancer Society, the Göran Gustafsson foundation, the Knut och Alice Wallenberg foundation, the EU integrated projects E-MEP and EICOSANOX. This Letter reflects only the authors’ views and the European Commission is not liable for any use that may be made of the information herein. We also thank staff at ESRF, BESSY, SLS and MAXLAB for beam time and assistance during data collection. We are grateful to T. Cornvik for help with artwork and to T. Bergman for help with mass spectrometry analysis.

Author Contributions A.W., T.H. and J.Z.H. cloned, expressed and purified LTC4 synthase. Further purification and characterization of LTC4 synthase in various detergents was performed by A.W., D.M.M. and E.O. and was supervised by S.E. D.M.M. crystallized LTC4 synthase and solved the structure together with A.A.M. and A.K. Crystallographic work was performed by D.M.M. under supervision of A.K. and P.N. Extensive crystal and derivative screening was conducted by D.M.M., A.K. and D.N. Overall project management and strategy was performed by J.Z.H. and P.N. Writing of the manuscript was performed jointly by P.N., J.Z.H., S.E. and D.M.M. All authors discussed the results and commented on the manuscript.

Coordinates and structure factors for LTC4 synthase in its apo form and in complex with GSH have been deposited in the Protein Data Bank with accession codes 2UUI and 2UUH, respectively.

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Authors and Affiliations

  1. Division of Biophysics,,

    Daniel Martinez Molina, Andreas Kohl, Damian Niegowski, Said Eshaghi & Pär Nordlund

  2. Division of Chemistry 2, Department of Medical Biochemistry and Biophysics,

    Anders Wetterholm, Eva Ohlson, Tove Hammarberg & Jesper Z. Haeggström

  3. Structural Genomics Consortium, Karolinska Institutet, 17177 Stockholm, Sweden,

    Pär Nordlund

  4. Department of Biochemistry and Biophysics, Stockholm University, S-106 91 Stockholm, Sweden,

    Daniel Martinez Molina & Damian Niegowski

  5. European Molecular Biology Laboratory, Grenoble outstation, 6 Rue Jules Horowitz, 38042 Grenoble Cedex 9, France,

    Andrew A. McCarthy

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Correspondence to Said Eshaghi, Jesper Z. Haeggström or Pär Nordlund.

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Supplementary information

Supplementary Information

This file contains Supplementary Figures S1–S5 and Supplementary Table S1. The Supplementary Figures illustrate a schematic drawing of the reaction catalyzed by LTC4 synthase, the metal coordination and crystal packing, a stereo view of the bound substrate, a lig plot diagram of the bound substrate and finally a structure based sequence alignment of the MAPEG family. A supplementary Table is included with crystallographic statistics. (PDF 1645 kb)

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Molina, D., Wetterholm, A., Kohl, A. et al. Structural basis for synthesis of inflammatory mediators by human leukotriene C4 synthase. Nature 448, 613–616 (2007). https://doi.org/10.1038/nature06009

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