Liposome-based measurement of light-driven chloride transport kinetics of halorhodopsin

Hasin Feroz; Bryan Ferlez; Hyeonji Oh; Hossein Mohammadiarani; Tingwei Ren; Carol S. Baker; John P. Gajewski; Daniel J. Lugar; Sandeep B. Gaudana; Peter Butler; Jonas Hühn; Matthias Lamping; Wolfgang J. Parak; Michael R. Blatt; Cheryl A. Kerfeld; Nicholas Smirnoff; Harish Vashisth; John H. Golbeck; Manish Kumar

doi:10.1016/j.bbamem.2021.183637

Liposome-based measurement of light-driven chloride transport kinetics of halorhodopsin

Hasin Feroz, Bryan Ferlez, Hyeonji Oh, Hossein Mohammadiarani, Tingwei Ren, Carol S. Baker, John P. Gajewski, Daniel J. Lugar, Sandeep B. Gaudana, Peter Butler, Jonas Hühn, Matthias Lamping, Wolfgang J. Parak, Michael R. Blatt, Cheryl A. Kerfeld, Nicholas Smirnoff, Harish Vashisth, John H. Golbeck, Manish Kumar

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3 Scopus citations

Abstract

We report a simple and direct fluorimetric vesicle-based method for measuring the transport rate of the light-driven ions pumps as specifically applied to the chloride pump, halorhodopsin, from Natronomonas pharaonis (pHR). Previous measurements were cell-based and methods to determine average single channel permeability challenging. We used a water-in-oil emulsion method for directional pHR reconstitution into two different types of vesicles: lipid vesicles and asymmetric lipid-block copolymer vesicles. We then used stopped-flow experiments combined with fluorescence correlation spectroscopy to determine per protein Cl- transport rates. We obtained a Cl⁻ transport rate of 442 (±17.7) Cl⁻/protein/s in egg phosphatidyl choline (PC) lipid vesicles and 413 (±26) Cl⁻/protein/s in hybrid block copolymer/lipid (BCP/PC) vesicles with polybutadine-polyethylene oxide (PB₁₂PEO₈) on the outer leaflet and PC in the inner leaflet at a photon flux of 1450 photons/protein/s. Normalizing to a per photon basis, this corresponds to 0.30 (±0.07) Cl⁻/photon and 0.28 (±0.04) Cl⁻/photon for pure PC and BCP/PC hybrid vesicles respectively, both of which are in agreement with recently reported turnover of ~500 Cl⁻/protein/s from flash photolysis experiments and with voltage-clamp measurements of 0.35 (±0.16) Cl⁻/photon in pHR-expressing oocytes as well as with a pHR quantum efficiency of ~30%.

Original language	English (US)
Article number	183637
Journal	Biochimica et Biophysica Acta - Biomembranes
Volume	1863
Issue number	8
DOIs	https://doi.org/10.1016/j.bbamem.2021.183637
State	Published - Aug 1 2021

All Science Journal Classification (ASJC) codes

Biophysics
Biochemistry
Cell Biology

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10.1016/j.bbamem.2021.183637

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Feroz, H., Ferlez, B., Oh, H., Mohammadiarani, H., Ren, T., Baker, C. S., Gajewski, J. P., Lugar, D. J., Gaudana, S. B., Butler, P., Hühn, J., Lamping, M., Parak, W. J., Blatt, M. R., Kerfeld, C. A., Smirnoff, N., Vashisth, H., Golbeck, J. H., & Kumar, M. (2021). Liposome-based measurement of light-driven chloride transport kinetics of halorhodopsin. Biochimica et Biophysica Acta - Biomembranes, 1863(8), Article 183637. https://doi.org/10.1016/j.bbamem.2021.183637

@article{2dcd43be7852496eb7bd1707ca923bdf,

title = "Liposome-based measurement of light-driven chloride transport kinetics of halorhodopsin",

abstract = "We report a simple and direct fluorimetric vesicle-based method for measuring the transport rate of the light-driven ions pumps as specifically applied to the chloride pump, halorhodopsin, from Natronomonas pharaonis (pHR). Previous measurements were cell-based and methods to determine average single channel permeability challenging. We used a water-in-oil emulsion method for directional pHR reconstitution into two different types of vesicles: lipid vesicles and asymmetric lipid-block copolymer vesicles. We then used stopped-flow experiments combined with fluorescence correlation spectroscopy to determine per protein Cl- transport rates. We obtained a Cl− transport rate of 442 (±17.7) Cl−/protein/s in egg phosphatidyl choline (PC) lipid vesicles and 413 (±26) Cl−/protein/s in hybrid block copolymer/lipid (BCP/PC) vesicles with polybutadine-polyethylene oxide (PB12PEO8) on the outer leaflet and PC in the inner leaflet at a photon flux of 1450 photons/protein/s. Normalizing to a per photon basis, this corresponds to 0.30 (±0.07) Cl−/photon and 0.28 (±0.04) Cl−/photon for pure PC and BCP/PC hybrid vesicles respectively, both of which are in agreement with recently reported turnover of ~500 Cl−/protein/s from flash photolysis experiments and with voltage-clamp measurements of 0.35 (±0.16) Cl−/photon in pHR-expressing oocytes as well as with a pHR quantum efficiency of ~30%.",

author = "Hasin Feroz and Bryan Ferlez and Hyeonji Oh and Hossein Mohammadiarani and Tingwei Ren and Baker, {Carol S.} and Gajewski, {John P.} and Lugar, {Daniel J.} and Gaudana, {Sandeep B.} and Peter Butler and Jonas H{\"u}hn and Matthias Lamping and Parak, {Wolfgang J.} and Blatt, {Michael R.} and Kerfeld, {Cheryl A.} and Nicholas Smirnoff and Harish Vashisth and Golbeck, {John H.} and Manish Kumar",

note = "Publisher Copyright: {\textcopyright} 2021",

year = "2021",

month = aug,

day = "1",

doi = "10.1016/j.bbamem.2021.183637",

language = "English (US)",

volume = "1863",

journal = "Biochimica et Biophysica Acta - Biomembranes",

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Feroz, H, Ferlez, B, Oh, H, Mohammadiarani, H, Ren, T, Baker, CS, Gajewski, JP, Lugar, DJ, Gaudana, SB, Butler, P, Hühn, J, Lamping, M, Parak, WJ, Blatt, MR, Kerfeld, CA, Smirnoff, N, Vashisth, H, Golbeck, JH & Kumar, M 2021, 'Liposome-based measurement of light-driven chloride transport kinetics of halorhodopsin', Biochimica et Biophysica Acta - Biomembranes, vol. 1863, no. 8, 183637. https://doi.org/10.1016/j.bbamem.2021.183637

TY - JOUR

T1 - Liposome-based measurement of light-driven chloride transport kinetics of halorhodopsin

AU - Feroz, Hasin

AU - Ferlez, Bryan

AU - Oh, Hyeonji

AU - Mohammadiarani, Hossein

AU - Ren, Tingwei

AU - Baker, Carol S.

AU - Gajewski, John P.

AU - Lugar, Daniel J.

AU - Gaudana, Sandeep B.

AU - Butler, Peter

AU - Hühn, Jonas

AU - Lamping, Matthias

AU - Parak, Wolfgang J.

AU - Blatt, Michael R.

AU - Kerfeld, Cheryl A.

AU - Smirnoff, Nicholas

AU - Vashisth, Harish

AU - Golbeck, John H.

AU - Kumar, Manish

PY - 2021/8/1

Y1 - 2021/8/1

N2 - We report a simple and direct fluorimetric vesicle-based method for measuring the transport rate of the light-driven ions pumps as specifically applied to the chloride pump, halorhodopsin, from Natronomonas pharaonis (pHR). Previous measurements were cell-based and methods to determine average single channel permeability challenging. We used a water-in-oil emulsion method for directional pHR reconstitution into two different types of vesicles: lipid vesicles and asymmetric lipid-block copolymer vesicles. We then used stopped-flow experiments combined with fluorescence correlation spectroscopy to determine per protein Cl- transport rates. We obtained a Cl− transport rate of 442 (±17.7) Cl−/protein/s in egg phosphatidyl choline (PC) lipid vesicles and 413 (±26) Cl−/protein/s in hybrid block copolymer/lipid (BCP/PC) vesicles with polybutadine-polyethylene oxide (PB12PEO8) on the outer leaflet and PC in the inner leaflet at a photon flux of 1450 photons/protein/s. Normalizing to a per photon basis, this corresponds to 0.30 (±0.07) Cl−/photon and 0.28 (±0.04) Cl−/photon for pure PC and BCP/PC hybrid vesicles respectively, both of which are in agreement with recently reported turnover of ~500 Cl−/protein/s from flash photolysis experiments and with voltage-clamp measurements of 0.35 (±0.16) Cl−/photon in pHR-expressing oocytes as well as with a pHR quantum efficiency of ~30%.

AB - We report a simple and direct fluorimetric vesicle-based method for measuring the transport rate of the light-driven ions pumps as specifically applied to the chloride pump, halorhodopsin, from Natronomonas pharaonis (pHR). Previous measurements were cell-based and methods to determine average single channel permeability challenging. We used a water-in-oil emulsion method for directional pHR reconstitution into two different types of vesicles: lipid vesicles and asymmetric lipid-block copolymer vesicles. We then used stopped-flow experiments combined with fluorescence correlation spectroscopy to determine per protein Cl- transport rates. We obtained a Cl− transport rate of 442 (±17.7) Cl−/protein/s in egg phosphatidyl choline (PC) lipid vesicles and 413 (±26) Cl−/protein/s in hybrid block copolymer/lipid (BCP/PC) vesicles with polybutadine-polyethylene oxide (PB12PEO8) on the outer leaflet and PC in the inner leaflet at a photon flux of 1450 photons/protein/s. Normalizing to a per photon basis, this corresponds to 0.30 (±0.07) Cl−/photon and 0.28 (±0.04) Cl−/photon for pure PC and BCP/PC hybrid vesicles respectively, both of which are in agreement with recently reported turnover of ~500 Cl−/protein/s from flash photolysis experiments and with voltage-clamp measurements of 0.35 (±0.16) Cl−/photon in pHR-expressing oocytes as well as with a pHR quantum efficiency of ~30%.

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U2 - 10.1016/j.bbamem.2021.183637

DO - 10.1016/j.bbamem.2021.183637

M3 - Article

C2 - 33930372

AN - SCOPUS:85105031460

SN - 0005-2736

VL - 1863

JO - Biochimica et Biophysica Acta - Biomembranes

JF - Biochimica et Biophysica Acta - Biomembranes

IS - 8

M1 - 183637

ER -

Liposome-based measurement of light-driven chloride transport kinetics of halorhodopsin

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