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Visualization of ATP Synthase Dimers in Mitochondria by Electron Cryo-tomography
10:39

Visualization of ATP Synthase Dimers in Mitochondria by Electron Cryo-tomography

Published on: September 14, 2014

Structure of intramembrane particles in proteoliposomes containing yeast mitochondrial ATPase

R D Lang, E Brown

    European Journal of Cell Biology
    |December 1, 1981
    PubMed

    Insights

    Reconstituted yeast mitochondrial oligomycin-sensitive ATPase forms larger vesicles and intramembrane particles (IMPs) in liposomes. These IMPs, potentially ATPase oligomers, exhibit hexagonal arrays at higher protein concentrations.

    Area of Science:

    • Biochemistry
    • Molecular Biology
    • Membrane Biophysics

    Background:

    • Mitochondrial ATPases are crucial for cellular energy production.
    • Reconstitution of membrane proteins into liposomes is essential for studying their structure and function.
    • Oligomycin-sensitive ATPase (OS-ATPase) from yeast mitochondria is a key enzyme in energy metabolism.

    Purpose of the Study:

    • To investigate the structural properties of reconstituted yeast mitochondrial OS-ATPase within phospholipid vesicles.
    • To determine the impact of OS-ATPase incorporation on liposome morphology and membrane structure.
    • To analyze the arrangement and dimensions of intramembrane particles (IMPs) formed by the reconstituted ATPase.

    Main Methods:

    • Isolation of OS-ATPase from yeast mitochondria.
    • Reconstitution of purified OS-ATPase into phospholipid vesicles using cholate dialysis.
    • Structural analysis of liposomes using freeze-fracture electron microscopy (FFEM).
    • Image processing of FFEM data for IMP characterization and array analysis.

    Main Results:

    • Reconstitution of OS-ATPase into liposomes resulted in larger vesicles (up to 500 nm) compared to empty liposomes (20-40 nm).
    • Incorporated ATPase formed IMPs on vesicle fracture faces, with frequency dependent on protein-to-lipid ratio.
    • IMPs exhibited a mean diameter of approximately 12.9 nm, and at high protein concentrations, formed hexagonal arrays with 13.7 nm spacing.

    Conclusions:

    • Yeast mitochondrial OS-ATPase can be successfully reconstituted into functional liposomes.
    • The reconstituted ATPase forms IMPs and influences liposome size, indicating its membrane integration.
    • Observed hexagonal arrays suggest that IMPs represent oligomeric forms of the OS-ATPase.

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