Functional proteomics protocol for the identification of interaction partners in Tetrahymena thermophila

Syed Nabeel-Shah; Jyoti Garg; Pata-Eting Kougnassoukou Tchara; Ronald E. Pearlman; Jean-Philippe Lambert; Jeffrey Fillingham

STAR Protocols (Mar 2021)

Functional proteomics protocol for the identification of interaction partners in Tetrahymena thermophila

Syed Nabeel-Shah,
Jyoti Garg,
Pata-Eting Kougnassoukou Tchara,
Ronald E. Pearlman,
Jean-Philippe Lambert,
Jeffrey Fillingham

Affiliations

Syed Nabeel-Shah: Department of Chemistry and Biology, Ryerson University, 350 Victoria Street, Toronto, ON M5B 2K3, Canada; Donnelly Centre, University of Toronto, Toronto, ON, M5S 3E1, Canada; Department of Molecular Genetics, University of Toronto, Toronto, ON, M5S 1A8, Canada; Corresponding author
Jyoti Garg: Department of Chemistry and Biology, Ryerson University, 350 Victoria Street, Toronto, ON M5B 2K3, Canada; Department of Biology, York University, 4700 Keele Street, Toronto, ON, M3J 1P3, Canada
Pata-Eting Kougnassoukou Tchara: Department of Molecular Medicine, Cancer Research Center, Big Data Research Center, Université Laval, Quebec City, QC, Canada; CHU de Québec Research Center, CHUL, 2705 Laurier Boulevard, Quebec City, QC, G1V 4G2, Canada
Ronald E. Pearlman: Department of Biology, York University, 4700 Keele Street, Toronto, ON, M3J 1P3, Canada
Jean-Philippe Lambert: Department of Molecular Medicine, Cancer Research Center, Big Data Research Center, Université Laval, Quebec City, QC, Canada; CHU de Québec Research Center, CHUL, 2705 Laurier Boulevard, Quebec City, QC, G1V 4G2, Canada
Jeffrey Fillingham: Department of Chemistry and Biology, Ryerson University, 350 Victoria Street, Toronto, ON M5B 2K3, Canada; Corresponding author

Journal volume & issue: Vol. 2, no. 1
p. 100362

Abstract

Read online

Summary: We describe an optimized protocol for one-step affinity purification of FZZ-tagged proteins followed by mass spectrometry analysis for the identification of protein-protein interactions in the ciliate protozoan Tetrahymena thermophila. The FZZ epitope tag contains 2 protein A moieties (ZZ) and a 3xFLAG separated by a TEV cleavage site, which can also be employed in tandem affinity purification. This protocol is versatile and is suitable to use for other common epitope tags and can be adapted for other ciliates.For complete details on the use and execution of this protocol, please refer to Garg et al. (2019).

Published in STAR Protocols

ISSN: 2666-1667 (Online)
Publisher: Elsevier
Country of publisher: United States
LCC subjects: Science: Science (General)
Website: https://star-protocols.cell.com/

About the journal

Abstract

Keywords