bims-tricox Biomed News
on Translation, ribosomes and COX
Issue of 2025–07–20
two papers selected by
Yash Verma, University of Zurich
  1. Structures of Chaetomium thermophilum TOM complexes with bound preproteins.
  2. Isolation of Mitochondria From Yeast to Estimate Mitochondrial Pools of Inorganic Phosphate.
  1. Proc Natl Acad Sci U S A. 2025 Jul 22. 122(29): e2507279122
    Structures of Chaetomium thermophilum TOM complexes with bound preproteins.
    Ahmed-Noor A Agip, Pamela Ornelas, Tzu-Jing Yang, Ermanno Uboldi, Sabine Häder, Melanie A McDowell, Werner Kühlbrandt.
      Mitochondria import most of their proteins from the cytoplasm through the TOM complex. Preproteins containing targeting signals are recognized by the TOM receptor subunits and translocated by Tom40 across the outer mitochondrial membrane. We present four structures of the preprotein-bound and preprotein-free TOM core and holo complexes from the thermophilic fungus Chaetomium thermophilum, obtained by single-particle electron cryomicroscopy. Our structures reveal the symmetric arrangement of two copies of the Tom20 receptor subunit in the TOM holo complex. Several different conformations of Tom20 within the TOM holo complex highlight the dynamic nature of the receptor. The structure of preprotein-bound Tom20 provides insight into the early stages of protein translocation.
    Keywords:  TOM complex; cryoEM; mitochondrial translocation; preproteins
    DOI:  https://doi.org/10.1073/pnas.2507279122
  2. Bio Protoc. 2025 Jul 05. 15(13): e5370
    Isolation of Mitochondria From Yeast to Estimate Mitochondrial Pools of Inorganic Phosphate.
    Swagata Adhikary, Vineeth Vengayil, Sunil Laxman.
      Mitochondria are dynamic organelles with essential roles in energetics and metabolism. Several metabolites are common to both the cytosolic and mitochondrial fractions of the cell. The compartmentalization of metabolites within the mitochondria allows specialized uses for mitochondrial metabolism. Inorganic phosphate (Pi) is one such critical metabolite required for ATP synthesis, via glycolysis and mitochondrial oxidative phosphorylation. Estimating total cellular Pi levels cannot distinguish the distribution of Pi pools across different cellular compartments, such as the cytosol and mitochondria, and therefore separate the contributions made toward glycolysis or other cytosolic metabolic processes vs. mitochondrial outputs. Quantifying Pi pools in mitochondria can therefore be very useful toward understanding mitochondrial metabolism and phosphate homeostasis. Here, we describe a protocol for the fairly rapid, efficient isolation of mitochondria from Saccharomyces cerevisiae by immunoprecipitation for quantitative estimation of mitochondrial and cytosolic Pi pools. This method utilizes magnetic beads to capture FLAG-tagged mitochondria (Tom20-FLAG) from homogenized cell lysates. This method provides a valuable tool to investigate changes in mitochondrial phosphate dynamics. Additionally, this protocol can be coupled with LC-MS approaches to quantitatively estimate mitochondrial metabolites and proteins and can be similarly used to assess other metabolite pools that are partitioned between the cytosol and mitochondria. Key features • This protocol describes how to isolate mitochondria from Saccharomyces cerevisiae for quantitative estimation of inorganic phosphate or other metabolites. • Mitochondria are efficiently isolated by immunoprecipitation using magnetic beads, bypassing the need for time-consuming density-based centrifugation. • This method can be integrated into LC-MS-based workflows to quantify mitochondrial metabolites and proteins.
    Keywords:  Differential centrifugation; Immunoprecipitation; Inorganic phosphate; Mitochondria isolation; Saccharomyces cerevisiae
    DOI:  https://doi.org/10.21769/BioProtoc.5370
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