Rapid flip-flop of phospholipids in endoplasmic reticulum membranes studied by a stopped-flow approach

Uwe Marx, G Lassmann, H G Holzhütter, D Wüstner, Peter Müller, A Höhlig, J Kubelt, Andreas Herrmann

Publikation: Bidrag til tidsskriftTidsskriftartikelForskningpeer review

Resumé

The transbilayer movement of short-chain spin-labeled and fluorescent 7-nitrobenz-2-oxa-1,3-diazol-4-yl (NBD) phospholipid analogs in rat liver microsomes is measured by stopped-flow mixing of labeled microsomes with bovine serum albumin (BSA) solution. Extraction of analogs from the outer leaflet of microsomes to BSA can be directly monitored in conjunction with electron paramagnetic resonance or fluorescence spectroscopy by taking advantage of the fact that the signal of spin-labeled or fluorescent analogs bound to BSA is different from that of the analogs inserted into membranes. From the signal kinetics, the transbilayer movement and the distribution of analogs in microsomal membranes can be derived provided the extraction of analogs by BSA is much faster in comparison to the transbilayer movement of analogs. Half-times of the back-exchange for spin-labeled and fluorescent analogs were <3.5 and <9.5 s, respectively. The unprecedented time resolution of the assay revealed that the transbilayer movement of spin-labeled analogs is much faster than previously reported. The half-time of the movement was about 16 s or even less at room temperature. Transmembrane movement of NBD-labeled analogs was six- to eightfold slower than that of spin-labeled analogs.

OriginalsprogEngelsk
TidsskriftBiophysical Journal
Vol/bind78
Udgave nummer5
Sider (fra-til)2628-40
Antal sider13
ISSN0006-3495
DOI
StatusUdgivet - maj 2000
Udgivet eksterntJa

Fingeraftryk

Bovine Serum Albumin
Phospholipids
Membranes
Electron Spin Resonance Spectroscopy
Liver Microsomes

Citer dette

Marx, Uwe ; Lassmann, G ; Holzhütter, H G ; Wüstner, D ; Müller, Peter ; Höhlig, A ; Kubelt, J ; Herrmann, Andreas. / Rapid flip-flop of phospholipids in endoplasmic reticulum membranes studied by a stopped-flow approach. I: Biophysical Journal. 2000 ; Bind 78, Nr. 5. s. 2628-40.
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abstract = "The transbilayer movement of short-chain spin-labeled and fluorescent 7-nitrobenz-2-oxa-1,3-diazol-4-yl (NBD) phospholipid analogs in rat liver microsomes is measured by stopped-flow mixing of labeled microsomes with bovine serum albumin (BSA) solution. Extraction of analogs from the outer leaflet of microsomes to BSA can be directly monitored in conjunction with electron paramagnetic resonance or fluorescence spectroscopy by taking advantage of the fact that the signal of spin-labeled or fluorescent analogs bound to BSA is different from that of the analogs inserted into membranes. From the signal kinetics, the transbilayer movement and the distribution of analogs in microsomal membranes can be derived provided the extraction of analogs by BSA is much faster in comparison to the transbilayer movement of analogs. Half-times of the back-exchange for spin-labeled and fluorescent analogs were <3.5 and <9.5 s, respectively. The unprecedented time resolution of the assay revealed that the transbilayer movement of spin-labeled analogs is much faster than previously reported. The half-time of the movement was about 16 s or even less at room temperature. Transmembrane movement of NBD-labeled analogs was six- to eightfold slower than that of spin-labeled analogs.",
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author = "Uwe Marx and G Lassmann and Holzh{\"u}tter, {H G} and D W{\"u}stner and Peter M{\"u}ller and A H{\"o}hlig and J Kubelt and Andreas Herrmann",
year = "2000",
month = "5",
doi = "10.1016/S0006-3495(00)76807-X",
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volume = "78",
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Marx, U, Lassmann, G, Holzhütter, HG, Wüstner, D, Müller, P, Höhlig, A, Kubelt, J & Herrmann, A 2000, 'Rapid flip-flop of phospholipids in endoplasmic reticulum membranes studied by a stopped-flow approach', Biophysical Journal, bind 78, nr. 5, s. 2628-40. https://doi.org/10.1016/S0006-3495(00)76807-X

Rapid flip-flop of phospholipids in endoplasmic reticulum membranes studied by a stopped-flow approach. / Marx, Uwe; Lassmann, G; Holzhütter, H G; Wüstner, D; Müller, Peter; Höhlig, A; Kubelt, J; Herrmann, Andreas.

I: Biophysical Journal, Bind 78, Nr. 5, 05.2000, s. 2628-40.

Publikation: Bidrag til tidsskriftTidsskriftartikelForskningpeer review

TY - JOUR

T1 - Rapid flip-flop of phospholipids in endoplasmic reticulum membranes studied by a stopped-flow approach

AU - Marx, Uwe

AU - Lassmann, G

AU - Holzhütter, H G

AU - Wüstner, D

AU - Müller, Peter

AU - Höhlig, A

AU - Kubelt, J

AU - Herrmann, Andreas

PY - 2000/5

Y1 - 2000/5

N2 - The transbilayer movement of short-chain spin-labeled and fluorescent 7-nitrobenz-2-oxa-1,3-diazol-4-yl (NBD) phospholipid analogs in rat liver microsomes is measured by stopped-flow mixing of labeled microsomes with bovine serum albumin (BSA) solution. Extraction of analogs from the outer leaflet of microsomes to BSA can be directly monitored in conjunction with electron paramagnetic resonance or fluorescence spectroscopy by taking advantage of the fact that the signal of spin-labeled or fluorescent analogs bound to BSA is different from that of the analogs inserted into membranes. From the signal kinetics, the transbilayer movement and the distribution of analogs in microsomal membranes can be derived provided the extraction of analogs by BSA is much faster in comparison to the transbilayer movement of analogs. Half-times of the back-exchange for spin-labeled and fluorescent analogs were <3.5 and <9.5 s, respectively. The unprecedented time resolution of the assay revealed that the transbilayer movement of spin-labeled analogs is much faster than previously reported. The half-time of the movement was about 16 s or even less at room temperature. Transmembrane movement of NBD-labeled analogs was six- to eightfold slower than that of spin-labeled analogs.

AB - The transbilayer movement of short-chain spin-labeled and fluorescent 7-nitrobenz-2-oxa-1,3-diazol-4-yl (NBD) phospholipid analogs in rat liver microsomes is measured by stopped-flow mixing of labeled microsomes with bovine serum albumin (BSA) solution. Extraction of analogs from the outer leaflet of microsomes to BSA can be directly monitored in conjunction with electron paramagnetic resonance or fluorescence spectroscopy by taking advantage of the fact that the signal of spin-labeled or fluorescent analogs bound to BSA is different from that of the analogs inserted into membranes. From the signal kinetics, the transbilayer movement and the distribution of analogs in microsomal membranes can be derived provided the extraction of analogs by BSA is much faster in comparison to the transbilayer movement of analogs. Half-times of the back-exchange for spin-labeled and fluorescent analogs were <3.5 and <9.5 s, respectively. The unprecedented time resolution of the assay revealed that the transbilayer movement of spin-labeled analogs is much faster than previously reported. The half-time of the movement was about 16 s or even less at room temperature. Transmembrane movement of NBD-labeled analogs was six- to eightfold slower than that of spin-labeled analogs.

KW - 4-Chloro-7-nitrobenzofurazan/analogs & derivatives

KW - Animals

KW - Biophysical Phenomena

KW - Biophysics

KW - Carrier Proteins/metabolism

KW - Cattle

KW - Electron Spin Resonance Spectroscopy

KW - Endoplasmic Reticulum/chemistry

KW - Fluorescent Dyes

KW - In Vitro Techniques

KW - Intracellular Membranes/chemistry

KW - Kinetics

KW - Membrane Lipids/chemistry

KW - Membrane Proteins/metabolism

KW - Microsomes, Liver/chemistry

KW - Models, Biological

KW - Phosphatidylcholines

KW - Phosphatidylethanolamines

KW - Phospholipid Transfer Proteins

KW - Phospholipids/chemistry

KW - Rats

KW - Serum Albumin, Bovine

KW - Spectrometry, Fluorescence

KW - Spin Labels

KW - Thermodynamics

U2 - 10.1016/S0006-3495(00)76807-X

DO - 10.1016/S0006-3495(00)76807-X

M3 - Journal article

C2 - 10777759

VL - 78

SP - 2628

EP - 2640

JO - Biophysical Journal

JF - Biophysical Journal

SN - 0006-3495

IS - 5

ER -