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Site-Directed Fluorescence Labeling (SDFL): TrIQ Methods Provide Insights Using the Fluorescent Probe Bimane

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Abstract

This review is in honor of Professor Wayne Hubbell’s 80th birthday, and his many contributions to the philosophy and practice of science. He continues to pioneer the use of site-directed spin labeling (SDSL) as a powerful tool for studying protein structure and dynamics. Inspired by his efforts, our lab has worked to develop site-directed fluorescence labeling (SDFL) as a complementary approach to SDSL. Throughout this process, we have emulated many of the novel methods and concepts Dr. Hubbell and his colleagues used during their development of SDSL. Some of our approaches are discussed here, not in a comprehensive or exhaustive way, but to highlight SDFL methods we have found to be robust, reproducible, and relatively easy to use. Specifically, we will focus on the development and application of SDFL using the fluorescent probe bimane, and how the unique properties of bimane can be exploited to glean information about protein structure and function. We also review past examples of how we have used bimane and SDFL to investigate the activation of G protein-coupled receptors (GPCRs) and their interaction with key binding partners.

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All data are contained within the manuscript and reference papers. Numerical values shown in the plots can be obtained upon request.

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Funding

This work was funded in part by R01 EY029343, R21 EY027959 and R21 DA043001 (DLF);  also T32 DA7262 and  F31 DA049438 (ADS).

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Authors and Affiliations

  1. Department of Chemical Physiology and Biochemistry, Oregon Health & Science University, 3181 SW Sam Jackson Park Rd, L224, MRB 504, Portland, OR, 97239, USA

    Anthony D. Shumate & David L. Farrens

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  1. Anthony D. Shumate
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  2. David L. Farrens
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Contributions

AS: conceptualization, methodology. Investigation: execution, validation and formal analysis of new unpublished data. Writing: original draft and writing, reviewing and editing. Visualization: generated figures. DLF: conceptualization, methodology. Writing: original draft and writing, reviewing and editing. Visualization: generated figures. Supervision, project administration, funding acquisition.

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Correspondence to David L. Farrens.

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Shumate, A.D., Farrens, D.L. Site-Directed Fluorescence Labeling (SDFL): TrIQ Methods Provide Insights Using the Fluorescent Probe Bimane. Appl Magn Reson 55, 101–126 (2024). https://doi.org/10.1007/s00723-023-01628-6

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