Light-driven chloride transport kinetics of halorhodopsin

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)

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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 > Institute of Molecular Cell and Systems Biology
Journal Name:Biophysical Journal
Publisher:Elsevier (Cell Press)
ISSN:0006-3495
ISSN (Online):1542-0086

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Project CodeAward NoProject NamePrincipal InvestigatorFunder's NameFunder RefLead Dept
563753MAGIC - A Multi-tiered approach to generating increased carbon dioxide in the chloroplastMichael BlattBiotechnology and Biological Sciences Research Council (BBSRC)BB/M01133X/1RI MOLECULAR CELL & SYSTEMS BIOLOGY
656891Analysing GORK clustering for enhanced stomatal controlMichael BlattBiotechnology and Biological Sciences Research Council (BBSRC)BB/M001601/1RI MOLECULAR CELL & SYSTEMS BIOLOGY