In vitro analysis of yeast mitochondrial protein import.
This unit describes methods for importing in vitro-translated or recombinant proteins into isolated yeast mitochondria and for exporting mitochondrial proteins translated in the yeast mitochondrial matrix into the inner mitochondrial membrane. The methods use mitochondria isolated from yeast cells and mitochondrial protein precursors derived from an in vitro transcription/translation reaction or purified from an E. coli recombinant protein expression system. The described translocation assays can be used to determine whether a protein is targeted to mitochondria and its location within the mitochondrion. It can also be used to study the import mechanism and to investigate mitochondrial matrix translation of proteins and their export.
Stuart
Rosemary A
RA
Marquette University, Milwaukee, Wisconsin, USA.
Koehler
Carla M
CM
D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D004926:Escherichia coli; D050259:Genes, Mitochondrial; D007202:Indicators and Reagents; D008928:Mitochondria / Q000378:metabolism* / Q000648:ultrastructure; D033681:Mitochondrial Membrane Transport Proteins / Q000378:metabolism; D051336:Mitochondrial Membranes / Q000378:metabolism; D024101:Mitochondrial Proteins / Q000235:genetics / Q000378:metabolism*; D014176:Protein Biosynthesis; D021382:Protein Sorting Signals / Q000502:physiology; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism / Q000648:ultrastructure*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*; D014158:Transcription, Genetic
D007202:Indicators and Reagents; D033681:Mitochondrial Membrane Transport Proteins; D024101:Mitochondrial Proteins; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins