By the same authors

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Validation of IMS-MS as a screening tool to identify type II kinase inhibitors of FGFR1 kinase

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Published copy (DOI)

Author(s)

  • Helen Beeston
  • Tobias Klein
  • Richard Norman
  • Julie Ann Tucker
  • Malcolm Anderson
  • Alison E. Ashcroft
  • Geoff Holdgate

Department/unit(s)

Publication details

JournalRapid Communications in Mass Spectrometry
DateSubmitted - 26 Apr 2021
DateAccepted/In press - 24 May 2021
DateE-pub ahead of print (current) - 26 May 2021
Early online date26/05/21
Original languageEnglish

Abstract

Rationale
The protein kinase FGFR1 regulates cellular processes in human development. As over-activity of FGFR1 is implicated with cancer, effective inhibitors are in demand. Type I inhibitors, which bind to the active form of FGFR1, are less effective than type II inhibitors, which bind to the inactive form. Screening to distinguish between type I and type II inhibitors is required.

Methods
X-Ray crystallography was used to indicate whether a range of potential inhibitors bind to the active or inactive FGFR1 kinase conformation. The binding affinity of each ligand to FGFR1 was measured using biochemical methods. ESI-IMS-MS in conjunction with collision-induced protein unfolding generated a conformational profile of each FGFR1-ligand complex. The results indicate that the protein’s conformational profile depends on whether the inhibitor is type I or type II.

Results
X-Ray crystallography confirmed which of the kinase inhibitors bind to the active or inactive form of FGFR1 kinase. Collision-induced unfolding combined with ESI-IMS-MS showed distinct differences in the FGFR1 folding landscape for type I and type II inhibitors. Biochemical studies indicated a similar range of FGFR1 affinities for both types of inhibitors, thus providing confidence that the conformational variations detected using ESI-IMS-MS can be interpreted unequivocally and that this is an effective screening method.

Conclusion
A robust ESI-IMS-MS method has been implemented to distinguish between the binding mode of type I and type II inhibitors by monitoring the conformational unfolding profile of FGFR1. This rapid method requires low sample concentrations and could be used as a high-throughput screening technique for the characterisation of novel kinase inhibitors.

Bibliographical note

© 2021 The Authors

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