Approaches to Analyze Protein-Protein Interactions of Membrane Proteins

Hunke, Sabine; Müller, Volker

doi:10.5772/38069

Cited by 5 publications

(10 citation statements)

References 121 publications

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“…Depending on the detergents used for permeabilization and solubilization of the membrane and subsequent purification protocol, one can get very different information about the protein:protein interactions within a particular membrane. In addition, the purification of native membrane proteins for biochemical studies requires large amounts of starting materials and highly specific antibodies raised against the membrane proteins involved in nanocluster formation [20] . Therefore, in recent years, live cell imaging techniques employing the confocal microscope and fluorescent protein-tagged (FP-tagged) proteins were frequently used to study the composition and dynamics of proteins on the membrane [21,22] .…”

Section: Methods To Study Nanoscale Membrane Organizationsmentioning

confidence: 99%

“…This makes the characterization of membrane complexes difficult. One way around this problem is to use chemical or photo-active crosslinkers prior to a detergent lysis [20] . For example, the Strep-tagged membrane protein interaction experiment (SPINE) uses formaldehyde to crosslink co-localized membrane proteins prior to their affinity purification with Streptavidin coupled beads [34,35] .…”

Section: Methods To Study Nanoscale Membrane Organizationsmentioning

confidence: 99%

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The nanoscale organization of the B lymphocyte membrane

Maity

Yang

Klaesener

et al. 2015

Biochimica et Biophysica Acta (BBA) - Molecular Cell Research

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The fluid mosaic model of Singer and Nicolson correctly predicted that the plasma membrane (PM) forms a lipid bi-layer containing many integral trans-membrane proteins. This model also suggested that most of these proteins were randomly dispersed and freely diffusing moieties. Initially, this view of a dynamic and rather unorganized membrane was supported by early observations of the cell surfaces using the light microscope. However, recent studies on the PM below the diffraction limit of visible light (~ 250 nm) revealed that, at nanoscale dimensions, membranes are highly organized and compartmentalized structures. Lymphocytes are particularly useful to study this nanoscale membrane organization because they grow as single cells and are not permanently engaged in cell:cell contacts within a tissue that can influence membrane organization. In this review, we describe the methods that can be used to better study the protein:protein interaction and nanoscale organization of lymphocyte membrane proteins, with a focus on the B cell antigen receptor (BCR). Furthermore, we discuss the factors that may generate and maintain these membrane structures.

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Section: Methods To Study Nanoscale Membrane Organizationsmentioning

confidence: 99%

Section: Methods To Study Nanoscale Membrane Organizationsmentioning

confidence: 99%

The nanoscale organization of the B lymphocyte membrane

Maity

Yang

Klaesener

et al. 2015

Biochimica et Biophysica Acta (BBA) - Molecular Cell Research

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“…As most part of the interactome is comprised of interactions among proteins, PPIs mediate many other types of interactions to bring them into a single interaction network. For instance, many transcription factors (in protein–DNA interactions) carry out their functions in joint with other proteins in a complex [2], and many receptors (in protein–ligand interactions) using PPIs in the downstream signalling cascades [3]. In theory, all PPIs and their properties can be precisely predicted from genomic information via computational approaches, however the practical accuracy and applicable scope are limited due to computational power and methodology.…”

Section: Introductionmentioning

confidence: 99%

In Vivo Analysis of Protein–Protein Interactions with Bioluminescence Resonance Energy Transfer (BRET): Progress and Prospects

Sun

Yang

Wang

et al. 2016

IJMS

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Proteins are the elementary machinery of life, and their functions are carried out mostly by molecular interactions. Among those interactions, protein–protein interactions (PPIs) are the most important as they participate in or mediate all essential biological processes. However, many common methods for PPI investigations are slightly unreliable and suffer from various limitations, especially in the studies of dynamic PPIs. To solve this problem, a method called Bioluminescence Resonance Energy Transfer (BRET) was developed about seventeen years ago. Since then, BRET has evolved into a whole class of methods that can be used to survey virtually any kinds of PPIs. Compared to many traditional methods, BRET is highly sensitive, reliable, easy to perform, and relatively inexpensive. However, most importantly, it can be done in vivo and allows the real-time monitoring of dynamic PPIs with the easily detectable light signal, which is extremely valuable for the PPI functional research. This review will take a comprehensive look at this powerful technique, including its principles, comparisons with other methods, experimental approaches, classifications, applications, early developments, recent progress, and prospects.

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“…The analysis of S-palmitoylated membrane protein-protein interactions in cells, however, can be challenging compared to soluble proteins. 3 Membrane protein complexes may only be maintained under a native and unique lipid environment, which is often destroyed during cell lysis and difficult to reconstitute in vitro . Moreover, the hydrophobic and amphiphilic nature of membrane proteins makes classical techniques such as co-immunoprecipitation, two-hybrid systems and native gel electrophoresis more difficult to implement compared to soluble proteins.…”

mentioning

confidence: 99%

“…Moreover, the hydrophobic and amphiphilic nature of membrane proteins makes classical techniques such as co-immunoprecipitation, two-hybrid systems and native gel electrophoresis more difficult to implement compared to soluble proteins. 3a These technical challenges are further exacerbated by the dynamic nature of S-palmitoylation. New tools are therefore needed to characterize S-palmitoylated membrane protein complexes in living cells for functional studies.…”

mentioning

confidence: 99%

Bifunctional Fatty Acid Chemical Reporter for Analyzing S-Palmitoylated Membrane Protein–Protein Interactions in Mammalian Cells

Peng

Hang

2015

J. Am. Chem. Soc.

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Studying the functions of S-palmitoylated proteins in cells can be challenging due to the membrane targeting property and dynamic nature of protein S-palmitoylation. New strategies are therefore needed to specifically capture S-palmitoylated protein complexes in cellular membranes for dissecting their functions in vivo. Here we present a bifunctional fatty acid chemical reporter, x-alk-16, which contains an alkyne and a diazirine, for metabolic labeling of S-palmitoylated proteins and photocrosslinking of their involved protein complexes in mammalian cells. We demonstrate that x-alk-16 can be metabolically incorporated into known S-palmitoylated proteins such as H-Ras and IFITM3, a potent antiviral protein, and induce covalent crosslinking of IFITM3 oligomerization as well as its specific interactions with other membrane proteins upon in-cell photoactivation. Moreover, integration of x-alk-16-induced photocrosslinking with label-free quantitative proteomics allows identification of new IFITM3 interacting proteins.

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Approaches to Analyze Protein-Protein Interactions of Membrane Proteins

Cited by 5 publications

References 121 publications

The nanoscale organization of the B lymphocyte membrane

The nanoscale organization of the B lymphocyte membrane

In Vivo Analysis of Protein–Protein Interactions with Bioluminescence Resonance Energy Transfer (BRET): Progress and Prospects

Bifunctional Fatty Acid Chemical Reporter for Analyzing S-Palmitoylated Membrane Protein–Protein Interactions in Mammalian Cells

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