Rapid-Scan Time-Resolved Gd-Gd Electron Paramagnetic Resonance (rs-TiGGER)

Proteins are dynamic molecular machines whose function depends on coordinated structural motion. While modern techniques such as cryo-electron microscopy and NMR have resolved static structures at high resolution, capturing these conformational changes in real time under biologically relevant conditions remains a central challenge in structural biology.

In the Sherwin Lab, we address this problem using site-directed spin labeling (SDSL) combined with high-field (8.6 T, 240 GHz) electron paramagnetic resonance (EPR) spectroscopy. This approach enables quantitative measurements of distance changes in doubly labeled proteins, providing direct access to nanoscale structural dynamics.

Using rapid-scan EPR with Gd³⁺ spin labeling (rs-TiGGER), we have directly observed light-induced conformational changes in the photoreceptor protein AsLOV2. Photoexcitation of its FMN chromophore initiates a cascade of structural changes, including undocking of the Jα helix on microsecond timescales, followed by relaxation back to the ground state over minutes. Measurements of the latter demonstrate the ability to track protein motion continuously (Figure 1).

Fig. 1: Price et al, 2024

This work is part of an interdisciplinary collaboration. The group of Prof. Songi Han (Northwestern University) provides complementary spin-labeling and magnetic resonance capabilities, including NMR and ODNP, enabling studies of pressure effects and hydration. At UC Santa Barbara, the group of Prof. Joan-Emma Shea contributes molecular dynamics simulations that connect measured distance changes to ensemble behavior and thermodynamic properties.

Ongoing efforts focus on extending rs-TiGGER capabilities through improved resonator design, including photonic bandgap structures to reduce dielectric loss and enhance time resolution. More broadly, we aim to reconstruct time-resolved “movies” of protein motion by combining measurements across multiple labeled sites. In the long term, such datasets may enable predictive models of protein dynamics analogous to structure prediction frameworks such as AlphaFold.

Principal Investigators

  • Prof. Mark Sherwin (Physics, UCSB)
  • Prof. Songi Han (Chemistry and Biochemistry, Northwestern)

Graduate Students

  • Johanna Schubert (Experimental Biophysics, UCSB)
  • Jackson Shepherd (Theoretical Biophysics, UCSB)
  • William Salvia (Biochem, Northwestern)

Collborators

  • Prof. Daniella Goldfarb (Weizmann Institute of Science)
  • Prof. Janet Lovett (University of St. Andrews)
  • Prof. Alex Smirnov (North Carolina State University)
  • Prof. David Parker, (Durham University)
  • Prof. Joan Emma-Shea (UCSB)
  • Prof. Max Wilson (UCSB)

Publications

Field-domain rapid-scan EPR at 240 GHz for studies of protein functional dynamics at room temperature
Brad D. Price, Antonín Sojka, Shiny Maity, I. Marcelo Chavez, Matthieu Starck, Maxwell Z. Wilson, Songi Han, Mark S. Sherwin
Journal of Magnetic Resonance 366, 107744 (2024)

Triggered functional dynamics of AsLOV2 by time-resolved electron paramagnetic resonance at high magnetic fields
Shiny Maity, Brad D. Price, C. Blake Wilson, Arnab Mukherjee, Matthieu Starck, David Parker, Maxwell Z. Wilson, Janet E. Lovett, Songi Han, Mark S. Sherwin
Angewandte Chemie International Edition 62(13), e202212832 (2023)

Quantitative analysis of zero-field splitting parameter distributions in Gd(III) complexes
Jessica A. Clayton, Katharina Keller, Mian Qi, Julia Wegner, Vanessa Koch, Henrik Hintz, Adelheid Godt, Songi Han, Gunnar Jeschke, Mark S. Sherwin, Maxim Yulikov
Physical Chemistry Chemical Physics 20, 10470 (2018)

Gd3+-Gd3+ Distances Exceeding 3nm Determined By Very High Frequency Continuous Wave Electron Paramagnetic Resonance
Jessica A. Clayton, Mian Qi, Adelheid Godt, Daniella Goldfarb, Songi Han, Mark S. Sherwin
Physical Chemistry Chemical Physics 19, 5127 (2017)

Determining The Oligomeric Structure Of Proteorhodopsin By Gd3+-Based Pulsed Dipolar Spectroscopy Of Multiple Distances
Devin T. Edwards, Thomas Huber, Sunyia Hussain, Katherine M. Stone, Maia Kinnebrew, Ilia Kaminker, Erez Matalon, Mark S. Sherwin, Daniella Goldfarb, Songi Han
Structure 22, 1677 (2014)

Extending The Distance Range Accessed With Continuous Wave EPR With Gd3+ Spin Probes At High Magnetic Fields
Devin T. Edwards, Zhidong Ma, Thomas J. Meade, Daniella Goldfarb, Songi Han, Mark S. Sherwin
Physical Chemistry Chemical Physics 15, 11313 (2013)

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