Single Vesicle Fluorescence-Bleaching Assay for Multi-Parameter Analysis of Proteoliposomes by Total Internal Reflection Fluorescence Microscopy

Research output: Contribution to journal › Journal article › Research › peer-review

Standard

Single Vesicle Fluorescence-Bleaching Assay for Multi-Parameter Analysis of Proteoliposomes by Total Internal Reflection Fluorescence Microscopy. / Veit, Sarina; Paweletz, Laura Charlotte; Bohr, Søren S-R; Menon, Anant K.; Hatzakis, Nikos S.; Günther-Pomorski, Thomas.

In: ACS applied materials & interfaces, Vol. 14, No. 26, 2022, p. 29659−29667.

Research output: Contribution to journal › Journal article › Research › peer-review

Harvard

Veit, S, Paweletz, LC, Bohr, SS-R, Menon, AK, Hatzakis, NS & Günther-Pomorski, T 2022, 'Single Vesicle Fluorescence-Bleaching Assay for Multi-Parameter Analysis of Proteoliposomes by Total Internal Reflection Fluorescence Microscopy', ACS applied materials & interfaces, vol. 14, no. 26, pp. 29659−29667. https://doi.org/10.1021/acsami.2c07454

APA

Veit, S., Paweletz, L. C., Bohr, S. S-R., Menon, A. K., Hatzakis, N. S., & Günther-Pomorski, T. (2022). Single Vesicle Fluorescence-Bleaching Assay for Multi-Parameter Analysis of Proteoliposomes by Total Internal Reflection Fluorescence Microscopy. ACS applied materials & interfaces, 14(26), 29659−29667. https://doi.org/10.1021/acsami.2c07454

Vancouver

Veit S, Paweletz LC, Bohr SS-R, Menon AK, Hatzakis NS, Günther-Pomorski T. Single Vesicle Fluorescence-Bleaching Assay for Multi-Parameter Analysis of Proteoliposomes by Total Internal Reflection Fluorescence Microscopy. ACS applied materials & interfaces. 2022;14(26):29659−29667. https://doi.org/10.1021/acsami.2c07454

Author

Veit, Sarina ; Paweletz, Laura Charlotte ; Bohr, Søren S-R ; Menon, Anant K. ; Hatzakis, Nikos S. ; Günther-Pomorski, Thomas. / Single Vesicle Fluorescence-Bleaching Assay for Multi-Parameter Analysis of Proteoliposomes by Total Internal Reflection Fluorescence Microscopy. In: ACS applied materials & interfaces. 2022 ; Vol. 14, No. 26. pp. 29659−29667.

Bibtex

@article{6671e9cd0ab34f9c9bfe24a477cb1144,
title = "Single Vesicle Fluorescence-Bleaching Assay for Multi-Parameter Analysis of Proteoliposomes by Total Internal Reflection Fluorescence Microscopy",
abstract = ": Reconstitution of membrane proteins into model membranes is an essential approach for their functional analysis under chemically defined conditions. Established model-membrane systems used in ensemble average measurements are limited by sample heterogeneity and insufficient knowledge of lipid and protein content at the single vesicle level, which limits quantitative analysis of vesicle properties and prevents their correlation with protein activity. Here, we describe a versatile total internal reflection fluorescence microscopy-based bleaching protocol that permits parallel analysis of multiple parameters (physical size, tightness, unilamellarity, membrane protein content, and orientation) of individual proteoliposomes prepared with fluorescently tagged membrane proteins and lipid markers. The approach makes use of commercially available fluorophores including the commonly used nitrobenzoxadiazole dye and may be applied to deduce functional molecular characteristics of many types of reconstituted fluorescently tagged membrane proteins.",
keywords = "ATPase, dithionite, image analysis, NBD-phospholipid, proteoliposome, single vesicle, TIRF microscopy, PLASMA-MEMBRANE ATPASE, SIZE DISTRIBUTIONS, RECONSTITUTION, PURIFICATION, LIPOSOMES, PROTEIN, QUANTIFICATION, RHODOPSIN, MECHANISM, STATE",
author = "Sarina Veit and Paweletz, {Laura Charlotte} and Bohr, {S{\o}ren S-R} and Menon, {Anant K.} and Hatzakis, {Nikos S.} and Thomas G{\"u}nther-Pomorski",
year = "2022",
doi = "10.1021/acsami.2c07454",
language = "English",
volume = "14",
pages = "29659−29667",
journal = "ACS applied materials & interfaces",
issn = "1944-8244",
publisher = "American Chemical Society",
number = "26",

}

RIS

TY - JOUR

T1 - Single Vesicle Fluorescence-Bleaching Assay for Multi-Parameter Analysis of Proteoliposomes by Total Internal Reflection Fluorescence Microscopy

AU - Veit, Sarina

AU - Paweletz, Laura Charlotte

AU - Bohr, Søren S-R

AU - Menon, Anant K.

AU - Hatzakis, Nikos S.

AU - Günther-Pomorski, Thomas

PY - 2022

Y1 - 2022

N2 - : Reconstitution of membrane proteins into model membranes is an essential approach for their functional analysis under chemically defined conditions. Established model-membrane systems used in ensemble average measurements are limited by sample heterogeneity and insufficient knowledge of lipid and protein content at the single vesicle level, which limits quantitative analysis of vesicle properties and prevents their correlation with protein activity. Here, we describe a versatile total internal reflection fluorescence microscopy-based bleaching protocol that permits parallel analysis of multiple parameters (physical size, tightness, unilamellarity, membrane protein content, and orientation) of individual proteoliposomes prepared with fluorescently tagged membrane proteins and lipid markers. The approach makes use of commercially available fluorophores including the commonly used nitrobenzoxadiazole dye and may be applied to deduce functional molecular characteristics of many types of reconstituted fluorescently tagged membrane proteins.

AB - : Reconstitution of membrane proteins into model membranes is an essential approach for their functional analysis under chemically defined conditions. Established model-membrane systems used in ensemble average measurements are limited by sample heterogeneity and insufficient knowledge of lipid and protein content at the single vesicle level, which limits quantitative analysis of vesicle properties and prevents their correlation with protein activity. Here, we describe a versatile total internal reflection fluorescence microscopy-based bleaching protocol that permits parallel analysis of multiple parameters (physical size, tightness, unilamellarity, membrane protein content, and orientation) of individual proteoliposomes prepared with fluorescently tagged membrane proteins and lipid markers. The approach makes use of commercially available fluorophores including the commonly used nitrobenzoxadiazole dye and may be applied to deduce functional molecular characteristics of many types of reconstituted fluorescently tagged membrane proteins.

KW - ATPase

KW - dithionite

KW - image analysis

KW - NBD-phospholipid

KW - proteoliposome

KW - single vesicle

KW - TIRF microscopy

KW - PLASMA-MEMBRANE ATPASE

KW - SIZE DISTRIBUTIONS

KW - RECONSTITUTION

KW - PURIFICATION

KW - LIPOSOMES

KW - PROTEIN

KW - QUANTIFICATION

KW - RHODOPSIN

KW - MECHANISM

KW - STATE

U2 - 10.1021/acsami.2c07454

DO - 10.1021/acsami.2c07454

M3 - Journal article

C2 - 35748880

VL - 14

SP - 29659−29667

JO - ACS applied materials & interfaces

JF - ACS applied materials & interfaces

SN - 1944-8244

IS - 26

ER -

ID: 312697920