A mitosome purification protocol based on percoll density gradients and its use in validating the mitosomal nature of Entamoeba histolytica mitochondrial Hsp70

Methods Mol Biol. 2007;390:167-77. doi: 10.1007/978-1-59745-466-7_11.

Abstract

Mitochondria are indispensable for aerobic respiration, but many microbial eukaryotes have lost this function through reductive evolution. Their modified mitochondria are known as hydrogenosomes or mitosomes depending on whether or not they produce molecular hydrogen. The intestinal parasite Entamoeba histolytica contains mitosomes whose role in cellular metabolism is unclear. Only three proteins have been shown thus far to reside in these organelles: the molecular chaperones Hsp10 and Hsp60 and an unusual ADP/ATP carrier. Here we describe the isolation of E. histolytica mitosomes by cellular fractionation and density gradient centrifugation and show that the mitochondrial-type chaperone Hsp70 is also housed in Entamoeba mitosomes.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Biochemistry / methods*
  • Cell Fractionation
  • Centrifugation, Density Gradient
  • Entamoeba histolytica / genetics
  • Entamoeba histolytica / metabolism*
  • HSP70 Heat-Shock Proteins / analysis*
  • Humans
  • Mitochondria / metabolism*
  • Organelles / metabolism*
  • Povidone / chemistry*
  • Silicon Dioxide / chemistry*

Substances

  • HSP70 Heat-Shock Proteins
  • Percoll
  • Silicon Dioxide
  • Povidone