Abstract：
:G protein-coupled receptors constitute one of the most important families of membrane receptors through which cells respond to extracellular stimuli. Receptors of this superfamily likely function as signal transduction complexes. The identification and analysis of their components provide new insights into a better understanding of these receptors' function and regulation. We used tandem-affinity purification and mass spectrometry as a systematic approach to characterize multiprotein complexes in the acetylcholine muscarinic receptor subfamily. To overcome the limitations associated with membrane protein receptor solubilization with detergents, we developed a strategy in which receptors are co-expressed with a cytoplasmic minigene construct, encoding the third intracellular loop and the C-terminal tail tagged to the tandem-affinity-cassette of each receptor subtype. Numerous protein complexes were identified, including many new interactions in various signalling pathways. Systematic identification data set together with protein interactions reported in the literature revealed a high degree of connectivity. These allow the proposal, for the first time, of an outline of the muscarinic interactome as a network of protein complexes and a context for a more reasoned and informed approach to drug discovery and muscarinic receptor subtype specificities.
journal_name
J Neurosci Methodsjournal_title
Journal of neuroscience methodsauthors
Borroto-Escuela DO,Correia PA,Romero-Fernandez W,Narvaez M,Fuxe K,Ciruela F,Garriga Pdoi
10.1016/j.jneumeth.2010.11.025subject
Has Abstractpub_date
2011-02-15 00:00:00pages
161-9issue
2eissn
0165-0270issn
1872-678Xpii
S0165-0270(10)00661-8journal_volume
195pub_type
杂志文章abstract：:Measurements of neurotransmitters in conscious animals have been restrictive in real-time. The use of specific enzymes within an amperometric probe, based upon a microdialysis membrane, may overcome some of these problems. I report the use of such a probe, with different perfusions of enzymes, to allow real-time measu...
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