Triggered Functional Dynamics of AsLOV2 by Time-Resolved Electron Paramagnetic Resonance at High Magnetic Fields |
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| Authors: | 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 |
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| Institution: | 1. Dept. of Chemistry and Biochemistry, Univ. of California, Santa Barbara, CA 93106 USA;2. Dept. of Physics, Univ. of California, Santa Barbara, CA 93106 USA;3. Dept. of Chemical Engineering, Univ. of California, Santa Barbara, CA 93106 USA;4. Dept. of Chemistry, Univ. of Durham, Durham, DH1 3LE UK;5. Dept. of Molecular, Cellular, and Developmental Biology, Univ. of California, Santa Barbara, CA 93106 USA;6. School of Physics and Astronomy and the Biomedical Sciences Research Complex, Univ. of St. Andrews, St. Andrews, KY16 9SS UK |
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| Abstract: | We present time-resolved Gd?Gd electron paramagnetic resonance (TiGGER) at 240 GHz for tracking inter-residue distances during a protein's mechanical cycle in the solution state. TiGGER makes use of Gd-sTPATCN spin labels, whose favorable qualities include a spin-7/2 EPR-active center, short linker, narrow intrinsic linewidth, and virtually no anisotropy at high fields (8.6 T) when compared to nitroxide spin labels. Using TiGGER, we determined that upon light activation, the C-terminus and N-terminus of AsLOV2 separate in less than 1 s and relax back to equilibrium with a time constant of approximately 60 s. TiGGER revealed that the light-activated long-range mechanical motion is slowed in the Q513A variant of AsLOV2 and is correlated to the similarly slowed relaxation of the optically excited chromophore as described in recent literature. TiGGER has the potential to valuably complement existing methods for the study of triggered functional dynamics in proteins. |
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| Keywords: | EPR Spectroscopy Gadolinium Protein Structures Tigger Time-Resolved Spectroscopy |
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