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Collapsin response mediator protein 2: high-resolution crystal structure sheds light on small-molecule binding, post-translational modifications, and conformational flexibility

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Abstract

Collapsin response mediator protein 2 (CRMP-2) is a neuronal protein involved in axonal pathfinding. Intense research is focusing on its role in various neurological diseases. Despite a wealth of studies, not much is known about the molecular mechanisms of CRMP-2 function in vivo. The detailed structure–function relationships of CRMP-2 have also largely remained unknown, in part due to the fact that the available crystal structures lack the C-terminal tail, which is known to be a target for many post-translational modifications and protein interactions. Although CRMP-2, and other CRMPs, belong to the dihydropyrimidinase family, they have lost the enzymatic active site. Drug candidates for CRMP-2-related processes have come up during the recent years, but no reports of CRMP-2 complexes with small molecules have emerged. Here, CRMP-2 was studied at 1.25-Å resolution using X-ray crystallography. In addition, ligands were docked into the homotetrameric structure, and the C-terminal tail of CRMP-2 was produced recombinantly and analyzed. We have obtained the human CRMP-2 crystal structure at atomic resolution and could identify small-molecule binding pockets in the protein. Structures obtained in different crystal forms highlight flexible regions near possible ligand-binding pockets. We also used the CRMP-2 structure to analyze known or suggested post-translational modifications at the 3D structural level. The high-resolution CRMP-2 structure was also used for docking experiments with the sulfur amino acid metabolite lanthionine ketimine and its ester. We show that the C-terminal tail is intrinsically disordered, but it has conserved segments that may act as interaction sites. Our data provide the most accurate structural data on CRMPs to date and will be useful in further computational and experimental studies on CRMP-2, its function, and its binding to small-molecule ligands.

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Acknowledgements

We wish to thank SGC Stockholm for providing the CRMP-2 cDNA, the Biocenter Oulu crystallization facility for excellent infrastructure, and EMBL-Hamburg/DESY for beamtime and outstanding beamline support. This research was funded by the Academy of Finland (PK), the Sigrid Jusélius Foundation (Finland) (PK), the Emil Aaltonen Foundation (Finland) (PK), Helse Vest (Bergen, Norway) (AB), and the Research and Science Foundation of the City of Hamburg (Germany) (PK). KH is funded for CRMP-2 research through the National Institutes of Health (NS082283 and NS093594).

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Authors and Affiliations

  1. Faculty of Biochemistry and Molecular Medicine and Biocenter Oulu, University of Oulu, Oulu, Finland

    Matti Myllykoski & Petri Kursula

  2. Department of Biomedicine, University of Bergen, Bergen, Norway

    Anne Baumann & Petri Kursula

  3. Division of Psychiatry, Haukeland University Hospital, Bergen, Norway

    Anne Baumann

  4. Arkansas College of Osteopathic Medicine, Fort Smith, AR, USA

    Kenneth Hensley

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  1. Matti Myllykoski
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Correspondence to Petri Kursula.

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KH is the inventor on US patent 7,683,055 (other patents pending) covering composition and use of LK derivatives including LKE and holds equity in a company engaged in commercial development of the technology.

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Myllykoski, M., Baumann, A., Hensley, K. et al. Collapsin response mediator protein 2: high-resolution crystal structure sheds light on small-molecule binding, post-translational modifications, and conformational flexibility. Amino Acids 49, 747–759 (2017). https://doi.org/10.1007/s00726-016-2376-z

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