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Molecular Physics
An International Journal at the Interface Between Chemistry and Physics
Volume 111, 2013 - Issue 18-19: Modern EPR Spectroscopy
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Invited Articles

Distance determination between low-spin ferric haem and nitroxide spin label using DEER: the neuroglobin case

, , , , , & show all
Pages 2855-2864 | Received 27 Mar 2013, Accepted 24 May 2013, Published online: 08 Jul 2013
 

Abstract

This work demonstrates for the first time the feasibility of using double electron–electron resonance (DEER) to determine the inter-spin distance between nitroxide spin labels and low-spin (S = 1/2) ferric haem centres. For these means, two human neuroglobin variants were spin labelled leading to singly labelled haem proteins with the nitroxide label on one of the natural Cys residues (Cys55 or Cys120). Room-temperature electron paramagnetic resonance was used to characterise the mobility of the nitroxide labels and X- and Q-band DEER experiments were performed to detect nitroxide–haem distances. Effects of residual nuclear modulations in the DEER traces were carefully evaluated. The DEER-derived distances were compared with theoretical predictions from an X-ray diffraction structure of human neuroglobin using a rotamer library approach as well as with distance information obtained from electron relaxation measurements. The structural biological implications of the spin-labelled side chains’ dynamics and of the obtained distances are also discussed.

Acknowledgements

S. Van Doorslaer acknowledges support from the Hercules Foundation, Flanders (contract AUHA013). S. Van Doorslaer and S. Dewilde also thank the Fund for Scientific Research – Flanders (FWO) and the University of Antwerp for funding.

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