FRAP beam-size analysis to measure palmitoylation-dependent membrane association dynamics and microdomain partitioning of Ras proteins

Yoav I. Henis; Barak Rotblat; Yoel Kloog

doi:10.1016/j.ymeth.2006.02.003

FRAP beam-size analysis to measure palmitoylation-dependent membrane association dynamics and microdomain partitioning of Ras proteins

Yoav I. Henis, Barak Rotblat, Yoel Kloog

Research output: Contribution to journal › Article › peer-review

48 Scopus citations

Abstract

Motions of membrane-associated proteins within and between membranes are essential for many cellular functions. We describe the application of fluorescence recovery after photobleaching (FRAP) beam-size analysis to investigate the role of palmitoylation in the membrane targeting and membrane association dynamics of H-Ras. The method described distinguishes between FRAP by lateral diffusion and by cytoplasmic exchange, and enables to obtain an estimate of the membrane affinity in live cells. These studies show distinct roles for the two palmitoylation sites (Cys181 and Cys184) on H-Ras, with different effects on membrane affinity and microlocalization.

Original language	English
Pages (from-to)	183-190
Number of pages	8
Journal	Methods
Volume	40
Issue number	2
DOIs	https://doi.org/10.1016/j.ymeth.2006.02.003
State	Published - 1 Oct 2006
Externally published	Yes

Keywords

Beam-size analysis
Cholesterol
Exchange
FRAP
H-Ras
Lateral diffusion
Membrane association
Palmitoylation

ASJC Scopus subject areas

Molecular Biology
General Biochemistry, Genetics and Molecular Biology

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10.1016/j.ymeth.2006.02.003

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title = "FRAP beam-size analysis to measure palmitoylation-dependent membrane association dynamics and microdomain partitioning of Ras proteins",

abstract = "Motions of membrane-associated proteins within and between membranes are essential for many cellular functions. We describe the application of fluorescence recovery after photobleaching (FRAP) beam-size analysis to investigate the role of palmitoylation in the membrane targeting and membrane association dynamics of H-Ras. The method described distinguishes between FRAP by lateral diffusion and by cytoplasmic exchange, and enables to obtain an estimate of the membrane affinity in live cells. These studies show distinct roles for the two palmitoylation sites (Cys181 and Cys184) on H-Ras, with different effects on membrane affinity and microlocalization.",

keywords = "Beam-size analysis, Cholesterol, Exchange, FRAP, H-Ras, Lateral diffusion, Membrane association, Palmitoylation",

author = "Henis, {Yoav I.} and Barak Rotblat and Yoel Kloog",

note = "Funding Information: This work was supported by the Wolfson Foundation and grant 339/02-3 from The Israel Science Foundation (YK). Y.I.H. is an incumbent of the Zalman Weinberg Chair in Cell Biology. Y.K. is an incumbent of the Jack H. Skirball Chair for Applied Neurobiology.",

year = "2006",

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doi = "10.1016/j.ymeth.2006.02.003",

language = "English",

volume = "40",

pages = "183--190",

journal = "Methods",

issn = "1046-2023",

publisher = "Academic Press Inc.",

number = "2",

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T1 - FRAP beam-size analysis to measure palmitoylation-dependent membrane association dynamics and microdomain partitioning of Ras proteins

AU - Henis, Yoav I.

AU - Rotblat, Barak

AU - Kloog, Yoel

N1 - Funding Information: This work was supported by the Wolfson Foundation and grant 339/02-3 from The Israel Science Foundation (YK). Y.I.H. is an incumbent of the Zalman Weinberg Chair in Cell Biology. Y.K. is an incumbent of the Jack H. Skirball Chair for Applied Neurobiology.

PY - 2006/10/1

Y1 - 2006/10/1

N2 - Motions of membrane-associated proteins within and between membranes are essential for many cellular functions. We describe the application of fluorescence recovery after photobleaching (FRAP) beam-size analysis to investigate the role of palmitoylation in the membrane targeting and membrane association dynamics of H-Ras. The method described distinguishes between FRAP by lateral diffusion and by cytoplasmic exchange, and enables to obtain an estimate of the membrane affinity in live cells. These studies show distinct roles for the two palmitoylation sites (Cys181 and Cys184) on H-Ras, with different effects on membrane affinity and microlocalization.

AB - Motions of membrane-associated proteins within and between membranes are essential for many cellular functions. We describe the application of fluorescence recovery after photobleaching (FRAP) beam-size analysis to investigate the role of palmitoylation in the membrane targeting and membrane association dynamics of H-Ras. The method described distinguishes between FRAP by lateral diffusion and by cytoplasmic exchange, and enables to obtain an estimate of the membrane affinity in live cells. These studies show distinct roles for the two palmitoylation sites (Cys181 and Cys184) on H-Ras, with different effects on membrane affinity and microlocalization.

KW - Beam-size analysis

KW - Cholesterol

KW - Exchange

KW - FRAP

KW - H-Ras

KW - Lateral diffusion

KW - Membrane association

KW - Palmitoylation

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U2 - 10.1016/j.ymeth.2006.02.003

DO - 10.1016/j.ymeth.2006.02.003

M3 - Article

C2 - 17012031

AN - SCOPUS:33748981090

SN - 1046-2023

VL - 40

SP - 183

EP - 190

JO - Methods

JF - Methods

IS - 2

ER -

FRAP beam-size analysis to measure palmitoylation-dependent membrane association dynamics and microdomain partitioning of Ras proteins

Abstract

Keywords

ASJC Scopus subject areas

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