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RESEARCH FRONT
Contents Vol 65(5)

Characterisation of Calmodulin Structural Transitions by Ion Mobility Mass Spectrometry

Antonio N. Calabrese A , Lauren A. Speechley A and Tara L. Pukala A B
+ Author Affiliations
- Author Affiliations

A School of Chemistry & Physics, The University of Adelaide, Adelaide, SA 5005, Australia.

B Corresponding author. Email: tara.pukala@adelaide.edu.au

Australian Journal of Chemistry 65(5) 504-511 https://doi.org/10.1071/CH12047
Submitted: 27 January 2012  Accepted: 7 April 2012   Published: 8 May 2012

Abstract

This study demonstrates the ability of travelling wave ion mobility-mass spectrometry to measure collision cross-sections of ions in the negative mode, using a calibration based approach. Here, negative mode ion mobility-mass spectrometry was utilised to understand structural transitions of calmodulin upon Ca2+ binding and complexation with model peptides melittin and the plasma membrane Ca2+ pump C20W peptide. Coexisting calmodulin conformers were distinguished on the basis of their mass and cross-section, and identified as relatively folded and unfolded populations, with good agreement in collision cross-section to known calmodulin geometries. Titration of calcium tartrate to physiologically relevant Ca2+ levels provided evidence for intermediately metalated species during the transition from apo- to holo-calmodulin, with collision cross-section measurements indicating that higher Ca2+ occupancy is correlated with more compact structures. The binding of two representative peptides which exemplify canonical compact (melittin) and extended (C20W) peptide-calmodulin binding models has also been interrogated by ion mobility mass spectrometry. Peptide binding to calmodulin involves intermediates with metalation states from 1–4 Ca2+, which demonstrate relatively collapsed structures, suggesting neither the existence of holo-calmodulin or a pre-folded calmodulin conformation is a prerequisite for binding target peptides or proteins. The biological importance of the different metal unsaturated calmodulin complexes, if any, is yet to be understood.


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