Purification of yeast peroxisomes.
Peroxisomes are ubiquitous subcellular organelles of eukaryotic cells. As with all organelles, peroxisomes can be purified from cell lysates using a combination of differential centrifugation and density gradient centrifugation. Here, we describe a method for purifying peroxisomes from the yeast Saccharomyces cerevisiae. The method involves gentle lysis of yeast spheroplasts, followed by differential centrifugation to obtain a crude organelle pellet enriched for peroxisomes and mitochondria. To separate peroxisomes from mitochondria, the organelle pellet is resuspended and spun through a sucrose density gradient. Peroxisomes purified in this way can be used to explore whether a protein of interest might be associated with the organelle.
Distel
Ben
B
Department of Medical Biochemistry, Academic Medical Center, University of Amsterdam, The Netherlands.
Kragt
Astrid
A
D002458:Cell Fractionation / Q000379:methods*; D002499:Centrifugation, Density Gradient; D007202:Indicators and Reagents; D008928:Mitochondria / Q000201:enzymology; D009172:Mycology / Q000379:methods; D020675:Peroxisomes / Q000737:chemistry* / Q000201:enzymology; D012441:Saccharomyces cerevisiae / Q000737:chemistry* / Q000201:enzymology / Q000648:ultrastructure
D007202:Indicators and Reagents