2005 Student Research Conference:
18th Annual Student Research Conference


Purification and Palmitoylation of Vac8p
Monica A. Stutz
Dr. Maurine E. Linder (Washington University) and Dr. Cynthia Cooper, Faculty Mentors

Palmitoylation is a post-translational lipid modification of many proteins involved in signal transduction, protein trafficking, and membrane association. The enzymes that catalyze this reaction are protein acyltransferases (PATs). In the yeast Saccharyomyces cerevisiae there is a family of seven putative PATs related by a common DHHC cysteine rich domain motif. Two of these PATs have known substrates. Vac8p, a yeast vacuolar protein, is myristoylated and palmitoylated at its N-terminus. The PAT responsible for this palmitoylation is unknown. The goal of this project was to purify myristoylated Vac8p from E. coli and use it as the substrate in an in vitro PAT assay to identify its PAT(s). Vac8p was partially purified using ion exchange chromatography. A subsequent PAT assay provided evidence that the DHHC-CRD protein Pfa3p, which also localizes to the yeast vacuole, palmitoylated Vac8p. Further experiments confirm the palmitoylation of Vac8p by Pfa3p.

Keywords: palmitoylation, Vac8p, DHHC-CRD, protein purification, yeast vacuole


Presentation Type: Oral Paper

Session: 10-3
Location: VH 1432
Time: 8:45

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