Abstract
Proteins of Saccharomyces cerevisiae can be metabolically labeled with (35) methionine. After labeling, a protocol is described for the mechanical disruption of yeast cells or conversion to spheroplasts, with subsequent lysis before immunoprecipitation of the proteins.
Materials and Reagents
Equipment
Procedure
Notes
Important notice for performing small-scale purification of solubilized membrane proteins from S35 labeled yeast extracts.
Important notice for running SDS-PAGE and Western Blot transfer with radioactive material.
Acknowledgments
This protocol has been modified and adapted in the Espenshade Lab, Johns Hopkins School of Medicine. Funding to support different projects that have used this protocol has come from NIH – National Heart, Lung, and Blood Institute, National Institute of Allergy and Infectious Diseases, the Pancreatic Cancer Action Network, and the American Heart Association.
References
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