Identification of Signaling Protein Complexes by Parallel Affinity Precipitation Coupled with Mass Spectrometry.
Protein-protein interactions play a pivotal role in both inter- and intra-cellular signaling. Identification of signaling protein complexes can thus shed important new insights into cell communications. We developed a parallel affinity precipitation protocol to overcome the disadvantages of the tandem affinity purification procedure, such as the potential disruption of target protein conformation, subcellular localization or function by epitope tags, the potential need of large amounts of cell culture or generation of stable cell lines, and relatively long duration the two-step precipitation takes. This new simplified assay of protein interaction is quick, economic and specific. This paper describes the details in the design and method of the assay.
Burnett School of Biomedical Science, College of Medicine, University of Central Florida, Orlando, FL, USA.
Heng
H
Lu
University at Albany Proteomics Facility, Center for Functional Genomics, Rensselaer, NY, USA.
Qishan
Q
Lin
Burnett School of Biomedical Science, College of Medicine, University of Central Florida, Orlando, FL, USA.
Jihe
J
Zhao
Myc tag; co-immunoprecipitation (co-IP); hemagglutinin tag; mass spectrometry; parallel affinity precipitation (PAP); protein–protein interaction; tandem affinity purification (TAP)