Mapping Protein–Protein Interactions Using Affinity Purification and Mass Spectrometry
Journal
Plant Genomics: Methods and Protocols
Journal Volume
1610
Pages
231 - 249
ISBN
978-1-4939-7001-8
Date Issued
2017
Author(s)
Abstract
The mapping of protein–protein interaction (PPI) networks and their dynamics are crucial steps to deciphering the function of a protein and its role in cellular pathways, making it critical to have comprehensive knowledge of a protein’s interactome. Advances in affinity purification and mass spectrometry technology (AP-MS) have provided a powerful and unbiased method to capture higher-order protein complexes and decipher dynamic PPIs. However, the unbiased calling of nonspecific interactions and the ability to detect transient interactions remains challenging when using AP-MS, thereby hampering the detection of biologically meaningful complexes. Additionally, there are plant-specific challenges with AP-MS, such as a lack of protein-specific antibodies, which must be overcome to successfully identify PPIs. Here we discuss and describe a protocol designed to bypass the traditional challenges of AP-MS and provide a roadmap to identify bona fide PPIs in plants.
Type
book part