Light-Driven Chloride Transport Kinetics of Halorhodopsin.

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.

journal_name

Biophys J

journal_title

Biophysical journal

authors

Feroz H,Ferlez B,Lefoulon C,Ren T,Baker CS,Gajewski JP,Lugar DJ,Gaudana SB,Butler PJ,Hühn J,Lamping M,Parak WJ,Hibberd JM,Kerfeld CA,Smirnoff N,Blatt MR,Golbeck JH,Kumar M

doi

10.1016/j.bpj.2018.06.009

subject

Has Abstract

pub_date

2018-07-17 00:00:00

pages

353-360

issue

2

eissn

0006-3495

issn

1542-0086

pii

S0006-3495(18)30685-4

journal_volume

115

pub_type

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