NDLI: Bipartite tetracysteine display requires site flexibility for ReAsH coordination

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Bipartite tetracysteine display requires site flexibility for ReAsH coordination

Content Provider	PubMed Central
Author	Goodman, Jessica L. Fried, Daniel B. Schepartz, Alanna
Abstract	Fluorescence is an essential tool for monitoring molecular interactions, cellular localization and pathway activation both in vitro and in vivo. The development of “pro-fluorescent” bisarsenical dyes in 1998 advanced this field in a significant way, as these dyes provide an alternative labeling strategy that complements some of the limitations of labeling with fluorescent proteins.[1] These small, cell permeable dyes, 4,5-bis(1,3,2-dithiarsolan-2-yl)fluorescein (FlAsH-EDT2, Figure S1A) and 4,5-bis(1,3,2-dithiarsolan-2-yl)resorufin (ReAsH-EDT2, Figure S1B) are non-fluorescent in the 1,2-ethanedithiol (EDT) bound form.[1, 2] Fluorescence occurs upon thiol-arsenic ligand exchange of EDT for a tetracysteine motif (Cys-Cys-Pro-Gly-Cys-Cys), permitting temporal fluorescent control.[1] Recently, we reported that the central Pro-Gly in the tetracysteine motif could be replaced with one or more folded proteins domains without a significant loss in bisarsenical affinity or brightness; we refer to this technique as ‘bipartite tetracysteine display’.[3] We showed, using a set of well characterized model systems, that bipartite tetracysteine display supports fluorescence-based differentiation between folded and unfolded protein states and can detect protein-protein interactions in live cells. The model constructs evaluated in our initial report shared two features in common: the dicysteine motifs were located near the protein N- and/or C-termini and the structures were primarily α-helical. Here we ask whether bipartite display also functions effectively within the context of two more complex protein architectures, p53 and EmGFP, in which the requisite Cys4 motif is introduced into loops and β-strands.
Related Links	http://dx.doi.org/10.1002/cbic.200900207
Ending Page	1647
Page Count	4
Starting Page	1644
File Format	PDF
ISSN	14394227
e-ISSN	14397633
Journal	Chembiochem : a European journal of chemical biology
Issue Number	10
Volume Number	10
Language	English
Publisher Date	2009-07-01
Access Restriction	Open
Subject Keyword	Medicine(all) Research in Higher Education
Content Type	Text
Resource Type	Article
Subject	Organic Chemistry Molecular Biology Biochemistry Molecular Medicine

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1	Ministry of Education (GoI), Department of Higher Education	Sanctioning Authority	https://www.education.gov.in/ict-initiatives
2	Indian Institute of Technology Kharagpur	Host Institute of the Project: The host institute of the project is responsible for providing infrastructure support and hosting the project	https://www.iitkgp.ac.in
3	National Digital Library of India Office, Indian Institute of Technology Kharagpur	The administrative and infrastructural headquarters of the project	Dr. B. Sutradhar bsutra@ndl.gov.in
4	Project PI / Joint PI	Principal Investigator and Joint Principal Investigators of the project	Dr. B. Sutradhar bsutra@ndl.gov.in Prof. Saswat Chakrabarti will be added soon
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