Pull-Down Assays
Generally, proteins function with partners or as components of large multi-protein complexes. Understanding these interactions is critical to investigating proteins’ biological pathways and cellular functions. Pull-down assay is a relatively easy, straightforward method to identify potential protein-binding partners. The method is also extensively used to confirm known interactions and to map interaction sites. The pull-down method relies on the immobilization of a fusion protein on beads that serves as a solid phase. The prey proteins binding to the bait protein can be captured and “pulled down” when the target-protein-binding beads are mixed and allowed to interact with the cell lysate. Prey proteins can come from multiple sources, including cell lysates, cell-free expression systems and extracted proteins. By subsequent washing and analysis using SDS-PAGE, Western Blot or Mass Spectrometry, a predicted interaction can be confirmed, and previously unknown interactions can be discovered.
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