1Michael Ford, 1Richard Jones, 1Ravi Amunugama, 1David Allen
1MS Bioworks, 3950 Varsity Drive, Ann Arbor, MI, U.S.A. 48108
2Jacqui Mendez, 2Nancy Murphy, 2Marie Schwinn, 2Hélène Benink, 2Danette L. Daniels, and 2Marjeta Urh
2Promega Corporation, 2800 Woods Hollow Road, Madison, WI U.S.A. 53711
The understanding of protein complex assembly and mapping of protein interactions has rapidly grown in recent years due to significant advances in mass spectrometry. As experiments turn toward characterization of the human proteome and the complexity it presents, a need remains to efficiently capture complexes intact, particularly weak or transient interactors as well as large multiprotein complexes. Here we present a new technology based upon the use of a protein fusion tag, termed HaloTag, which allows for highly specific and covalent immobilization of proteins complexes, as well as the ability to do correlative cellular localization studies using fluorescent ligands. In studies presented here, this technology was used to further understanding of the multi-protein Mediator complex assemblies and their cellular function.