Feroz, H. et al. (2018) Light-driven chloride transport kinetics of halorhodopsin. Biophysical Journal, 115(2), pp. 353-360. (doi: 10.1016/j.bpj.2018.06.009) (PMID:30021110) (PMCID:PMC6051019)
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Abstract
Despite growing interest in light-driven ion pumps for use in optogenetics, current estimates of their transport rates span two orders of magnitude due to challenges in measuring slow transport processes and determining protein concentration and/or orientation in membranes in vitro. In this study, we report, to our knowledge, the first direct quantitative measurement of light-driven Cl transport rates of the anion pump halorohodopsin from Natronomonas pharaonis (NpHR). We used light-interfaced voltage clamp measurements on NpHR-expressing oocytes to obtain a transport rate of 219 (± 98) Cl /protein/s for a photon flux of 630 photons/protein/s. The measurement is consistent with the literature-reported quantum efficiency of ∼30% for NpHR, i.e., 0.3 isomerizations per photon absorbed. To reconcile our measurements with an earlier-reported 20 ms rate-limiting step, or 35 turnovers/protein/s, we conducted, to our knowledge, novel consecutive single-turnover flash experiments that demonstrate that under continuous illumination, NpHR bypasses this step in the photocycle.
| Item Type: | Articles |
|---|---|
| Status: | Published |
| Refereed: | Yes |
| Glasgow Author(s) Enlighten ID: | Blatt, Professor Michael and Lefoulon, Dr Cecile |
| Authors: | Feroz, H., Ferlez, B., Lefoulon, C., Ren, T., Baker, C. S., Gajewski, J. P., Lugar, D. J., Gaudana, S. B., Butler, P. J., Hühn, J., Lamping, M., Parak, W. J., Hibberd, J. M., Kerfeld, C. A., Smirnoff, N., Blatt, M. R., Golbeck, J. H., and Kumar, M. |
| College/School: | College of Medical Veterinary and Life Sciences > School of Molecular Biosciences |
| Journal Name: | Biophysical Journal |
| Publisher: | Elsevier (Cell Press) |
| ISSN: | 0006-3495 |
| ISSN (Online): | 1542-0086 |
| Copyright Holders: | Copyright © 2018 Biophysical Society |
| First Published: | First published in Biophysical Journal 115(2):353-360 |
| Publisher Policy: | Reproduced in accordance with the copyright policy of the publisher |
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