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TitleHigh-Performance Intensiometric Direct- and Inverse-Response Genetically Encoded Biosensors for Citrate.
Publication TypeJournal Article
Year of Publication2020
AuthorsZhao, Yufeng, Yi Shen, Yurong Wen, and Robert E. Campbell
JournalACS Cent Sci
Volume6
Issue8
Pagination1441-1450
Date Published2020 Aug 26
ISSN2374-7943
Abstract

Motivated by the growing recognition of citrate as a central metabolite in a variety of biological processes associated with healthy and diseased cellular states, we have developed a series of high-performance genetically encoded citrate biosensors suitable for imaging of citrate concentrations in mammalian cells. The design of these biosensors was guided by structural studies of the citrate-responsive sensor histidine kinase and took advantage of the same conformational changes proposed to propagate from the binding domain to the catalytic domain. Following extensive engineering based on a combination of structure guided mutagenesis and directed evolution, we produced an inverse-response biosensor (Δ/ ≈ 18) designated Citroff1 and a direct-response biosensor (Δ/ ≈ 9) designated Citron1. We report the X-ray crystal structure of Citron1 and demonstrate the utility of both biosensors for qualitative and quantitative imaging of steady-state and pharmacologically perturbed citrate concentrations in live cells.

DOI10.1021/acscentsci.0c00518
Alternate JournalACS Cent Sci
PubMed ID32875085
PubMed Central IDPMC7453566