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Updated: Oct 19, 2025

Assessment of Submitochondrial Protein Localization in Budding Yeast Saccharomyces cerevisiae
Published on: July 19, 2021
Assessment of Mitochondrial Protein Topology and Membrane Insertion
Kerstin Schäfer1, Carina Engstler1, Korbinian Dischinger1
1Department Biologie I-Botanik, Ludwig-Maximilians-Universität München, Planegg-Martinsried, Germany.
Abstract:
Analyzing the membrane integrity and topology of a mitochondrial protein is essential for truly understanding its function. In this chapter, we demonstrate how to analyze mitochondrial membrane proteins using both an immunological-based assay and an in vivo self-assembling GFP approach. First, immunological approaches to investigate the solubility or membrane association of a protein within mitochondria are described. With this method, we demonstrate how the topology of soluble domains of a membrane-integrated protein can be determined. Using protein-specific antibodies, the localization of the soluble domains of a protein are analyzed by a proteolytic-cleavage approach using proteinase K in mitochondria, outer membrane-ruptured mitochondria, and solubilized mitochondrial membranes. In a second approach, we determine the topology of plant mitochondrial proteins using an in vivo self-assembling GFP localization approach.
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