Preprints
https://doi.org/10.5194/mr-2021-7
https://doi.org/10.5194/mr-2021-7

  05 Feb 2021

05 Feb 2021

Review status: a revised version of this preprint was accepted for the journal MR.

High Affinity Tamoxifen Analogues Retain Extensive Positional Disorder when Bound to Calmodulin

Lilia Milanesi1,2, Clare R. Trevitt1, Brian Whitehead1, Andrea M. Hounslow1, Salvador Tomas2, Laszlo L. P. Hosszu1,3, Christopher A. Hunter4, and Jonathan P. Waltho1,5 Lilia Milanesi et al.
  • 1Department of Molecular Biology and Biotechnology, University of Sheffield, Sheffield S10 2TN, UK
  • 2Department of Biological Sciences, School of Science, Birkbeck University of London, London WC1E 7HX, UK
  • 3Medical Research Council Prion Unit, University College of London Institute of Neurology, Queen Square, London WCN1 3BG, UK
  • 4Department of Chemistry, University of Cambridge, Lensfield Road, Cambridge CB2 1EW, UK
  • 5Manchester Institute of Biotechnology, University of Manchester, 131 Princess St, Manchester, M1 7DN, UK

Abstract. Using a combination of NMR and fluorescence measurements we have investigated the structure and dynamics of the complexes formed between calcium loaded calmodulin (CaM) and the potent breast cancer inhibitor idoxifene, a derivative of tamoxifen. High affinity binding (Kd ~ 300 nM) saturates with a 2:1 idoxifene:CaM complex. The complex is an ensemble where each idoxifene molecule is predominantly in the vicinity of one of the two hydrophobic patches of CaM but, in contrast with the lower affinity antagonists TFP, J-8 and W-7, does not substantially occupy the hydrophobic pocket. At least four idoxifene orientations per domain of CaM are necessary to satisfy the intermolecular NOE restraints, and this requires that the idoxifene molecules switch rapidly between positions. The CaM molecule is predominantly in the form where the N and C-terminal domains are in close proximity allowing for the idoxifene molecules to contact both domains simultaneously. Hence, the 2:1 idoxifene:CaM complex illustrates how high affinity binding occurs without the loss of extensive positional dynamics.

Lilia Milanesi et al.

Status: final response (author comments only)

Comment types: AC – author | RC – referee | CC – community | EC – editor | CEC – chief editor | : Report abuse
  • RC1: 'Comment on mr-2021-7', Walter Chazin, 23 Feb 2021
    • AC1: 'Reply on RC1', Jonathan Waltho, 23 Feb 2021
  • RC2: 'Comment on mr-2021-7', Anonymous Referee #2, 04 Mar 2021
    • AC2: 'Reply on RC2', Jonathan Waltho, 19 Mar 2021

Lilia Milanesi et al.

Lilia Milanesi et al.

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Short summary
The overall aim of the study is to provide a basis from which to improve the ability of tamoxifen family drugs to reduce the activity of a secondary target protein, calmodulin, during tumour development. The main conclusion is that the binding of a tamoxifen analogue is quite unlike that of other anti-calmodulin compounds, in that two drug molecules bring the two domains of calmodulin into close proximity, but they are not fixed in orientation relative to the protein.