Monitoring Complex Formation by Relaxation-Induced Pulse Electron Paramagnetic Resonance Distance Measurements.

Angeliki Giannoulis,Bela E. Bode,Maria Oranges

doi:10.1002/cphc.201700666

Abstract

Biomolecular complexes are often multimers fueling the demand for methods that allow unraveling their composition and geometric arrangement. Pulse electron paramagnetic resonance (EPR) spectroscopy is increasingly applied for retrieving geometric information on the nanometer scale. The emerging RIDME (relaxation‐induced dipolar modulation enhancement) technique offers improved sensitivity in distance experiments involving metal centers (e.g. on metalloproteins or proteins labelled with metal ions). Here, a mixture of a spin labelled ligand with increasing amounts of paramagnetic CuII ions allowed accurate quantification of ligand‐metal binding in the model complex formed. The distance measurement was highly accurate and critical aspects for identifying multimerization could be identified. The potential to quantify binding in addition to the high‐precision distance measurement will further increase the scope of EPR applications.

Highlights

Biomolecular complexes are often multimers fueling the demand for methods that allow unraveling their composition and geometric arrangement
Pulse electron paramagnetic resonance (EPR) has proven of utmost value for studying complex biological systems and revealing topology information not accessible by other methods
The impact of pulse EPR on structural research has sparked a renaissance of EPR methodology involving new hardware,[5] pulse sequences[6] and computational tools.[7]

Summary

Introduction

Biomolecular complexes are often multimers fueling the demand for methods that allow unraveling their composition and geometric arrangement. In PELDOR the number of electron spins per nano-object can be retrieved from the depth of the dipolar oscillations (D, Figure 1, panel A, left).[13] Assessing multimerization degrees in a similar fashion via relaxation-based pulse EPR would be a valuable addition to the distance information in the data. If the signal is the linear superposition of contributions from bound and unbound L, the depth of the dipolar modulations D will report the fraction of ligand bound to a metal ion.

Results

Conclusion