Characterization of membrane protein interactomes by Co-interacting Protein Identification Technology (CoPIT)
Finding interaction partners of proteins is an important step in understanding their function. While co-immunoprecipitation of proteins followed by mass spectrometric identification (AP-MS) has become a standard approach to identify interaction partners, characterization of membrane protein interactomes is still challenging because of the typically low abundance and high hydrophobicity of membrane proteins. Here, we describe CoPIT (for Co-interacting Protein Identification Technology), an approach to comprehensively identify and characterize the interactome of membrane proteins. The CoPIT protocol includes experimental and computational methods to establish membrane protein interactome networks and to monitor interactome changes upon perturbation. While the approach in particular improves membrane protein interactome analysis as demonstrated by using a low abundant membrane protein (Cystic Fibrosis Transmembrane Conductance Regulator (CFTR)) as a model, it is applicable to all types of proteins. The time required to complete the protocol varies from 3 days to several weeks depending on whether all or only part of it is carried out.
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Posted 30 Nov, 2015
Characterization of membrane protein interactomes by Co-interacting Protein Identification Technology (CoPIT)
Posted 30 Nov, 2015
Finding interaction partners of proteins is an important step in understanding their function. While co-immunoprecipitation of proteins followed by mass spectrometric identification (AP-MS) has become a standard approach to identify interaction partners, characterization of membrane protein interactomes is still challenging because of the typically low abundance and high hydrophobicity of membrane proteins. Here, we describe CoPIT (for Co-interacting Protein Identification Technology), an approach to comprehensively identify and characterize the interactome of membrane proteins. The CoPIT protocol includes experimental and computational methods to establish membrane protein interactome networks and to monitor interactome changes upon perturbation. While the approach in particular improves membrane protein interactome analysis as demonstrated by using a low abundant membrane protein (Cystic Fibrosis Transmembrane Conductance Regulator (CFTR)) as a model, it is applicable to all types of proteins. The time required to complete the protocol varies from 3 days to several weeks depending on whether all or only part of it is carried out.
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