Chromophore carbonyl twisting in fluorescent biosensors encodes direct readout of protein conformations with multicolor switching

Allert, Malin J.; Kumar, Shivesh; Wang, You; Beese, Lorena S.; Hellinga, Homme W.

doi:10.1038/s42004-023-00982-7

Chromophore carbonyl twisting in fluorescent biosensors encodes direct readout of protein conformations with multicolor switching

Journal Article · Sat Aug 19 00:00:00 EDT 2023 · Communications Chemistry

DOI:https://doi.org/10.1038/s42004-023-00982-7· OSTI ID:2423312

^[1]; Kumar, Shivesh ^[2]; Wang, You ^[1]; Beese, Lorena S. ^[1]; ^[1]

Duke Univ., Durham, NC (United States). Medical Center
Washington Univ., St. Louis, MO (United States); Duke Univ., Durham, NC (United States). Medical Center

Fluorescent labeling of proteins is a powerful tool for probing structure-function relationships with many biosensing applications. Structure-based rules for systematically designing fluorescent biosensors require understanding ligand-mediated fluorescent response mechanisms which can be challenging to establish. We installed thiol-reactive derivatives of the naphthalene-based fluorophore Prodan into bacterial periplasmic glucose-binding proteins. Glucose binding elicited paired color exchanges in the excited and ground states of these conjugates. X-ray structures and mutagenesis studies established that glucose-mediated color switching arises from steric interactions that couple protein conformational changes to twisting of the Prodan carbonyl relative to its naphthalene plane. Mutations of residues contacting the carbonyl can optimize color switching by altering fluorophore conformational equilibria in the apo and glucose-bound proteins. A commonly accepted view is that Prodan derivatives report on protein conformations via solvatochromic effects due to changes in the dielectric of their local environment. Here we show that instead Prodan carbonyl twisting controls color switching. These insights enable structure-based biosensor design by coupling ligand-mediated protein conformational changes to internal chromophore twists through specific steric interactions between fluorophore and protein.

View Accepted Manuscript (DOE)

Research Organization:: Argonne National Laboratory (ANL), Argonne, IL (United States). Advanced Photon Source (APS); Lawrence Berkeley National Laboratory (LBNL), Berkeley, CA (United States). Advanced Light Source (ALS)

Sponsoring Organization:: USDOE Office of Science (SC), Basic Energy Sciences (BES); National Institute of General Medical Sciences (NIGMS)

Grant/Contract Number:: AC02-06CH11357; AC02-05CH11231

OSTI ID:: 2423312

Journal Information:: Communications Chemistry, Journal Name: Communications Chemistry Journal Issue: 1 Vol. 6; ISSN 2399-3669

Publisher:: Springer NatureCopyright Statement

Country of Publication:: United States

Language:: English

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