Structural Requirements in the Transmembrane Domain of GLIC Revealed by Incorporation of Noncanonical Histidine Analogs
Abstract
The cyanobacterial pentameric ligand-gated ion channel GLIC, a homolog of the Cys-loop receptor superfamily, has provided useful structural and functional information about its eukaryotic counterparts. X-ray diffraction data and site-directed mutagenesis have previously implicated a transmembrane histidine residue (His234) as essential for channel function. Here, we investigated the role of His234 via synthesis and incorporation of histidine analogs and α-hydroxy acids using in vivo nonsense suppression. Receptors were expressed heterologously in Xenopus laevis oocytes, and whole-cell voltage-clamp electrophysiology was used to monitor channel activity. We show that an interhelix hydrogen bond involving His234 is important for stabilization of the open state, and that the shape and basicity of its side chain are highly sensitive to perturbations. In contrast, our data show that two other His residues are not involved in the acid-sensing mechanism.
Additional Information
© 2014 Elsevier Ltd. Received: July 25, 2014; Revised: September 23, 2014; Accepted: October 13, 2014; Published: December 18, 2014. This work was supported by grants from the NIH (NS034407 to D.A.D.) and the Wellcome Trust (WT 81925 to S.C.R.L., who is a Wellcome Trust Senior Research Fellow in Basic Biomedical Science) and NIH training grant NIH/ NRSA GM07616 to M.R. We would like to thank Fiona Love, Rell Parker, and Dr. Brandon Henderson for excellent technical assistance and Henry A. Lester and Mona Alqazzaz for helpful discussions.Attached Files
Accepted Version - nihms-645930.pdf
Supplemental Material - mmc1.pdf
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Additional details
- PMCID
- PMC4291181
- Eprint ID
- 53167
- DOI
- 10.1016/j.chembiol.2014.10.019
- Resolver ID
- CaltechAUTHORS:20150105-093951570
- NIH
- NS034407
- Wellcome Trust
- WT 81925
- NIH
- GM07616
- Created
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2015-01-06Created from EPrint's datestamp field
- Updated
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2021-11-10Created from EPrint's last_modified field