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Method to Visualize and Analyze Membrane Interacting Proteins by Transmission Electron Microscopy
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Published on: March 5, 2017

Lipid bilayer composition affects transmembrane protein orientation and function.

Katie D Hickey1, Mary M Buhr

  • 1Department of Animal and Poultry Science, University of Guelph, Guelph, ON, Canada N1G 2W1.

Journal of Lipids
|April 15, 2011
PubMed
Summary

Sperm membrane lipid composition influences protein orientation, crucial for fertilization. Altering lipids changes how Na(+) K(+)-ATPase is positioned within sperm membranes.

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Area of Science:

  • Biochemistry
  • Cell Biology
  • Reproductive Science

Background:

  • Sperm membranes undergo significant changes during capacitation, a process essential for fertilization.
  • Lipid composition and microdomains within sperm membranes are hypothesized to play a role in enabling capacitation.
  • Understanding the influence of the lipid environment on membrane proteins is key to understanding sperm function.

Purpose of the Study:

  • To investigate the effect of sperm membrane lipid composition on protein incorporation, function, and orientation.
  • To determine how altering lipid composition impacts the structure and function of specific membrane proteins like Na(+) K(+)-ATPase.

Main Methods:

  • Liposomes were constructed using lipids extracted from bull sperm membranes.
  • Proteins, including Na(+) K(+)-ATPase and α-amylase, were incorporated into liposomes with varying lipid compositions.
  • Fluorescence resonance energy transfer (FRET) was used to confirm protein inclusion and orientation.
  • Colourometric assays measured protein function via ATP cleavage and phosphate production.

Main Results:

  • Fluorescence resonance energy transfer confirmed the successful incorporation of proteins into the liposomes.
  • Protein function, specifically the ATPase activity of Na(+) K(+)-ATPase, was confirmed through phosphate production assays.
  • In liposomes made solely of native sperm lipids, Na(+) K(+)-ATPase (sodium-potassium adenosine triphosphatase) was oriented with its β subunit facing outward.
  • Modifying the lipid composition to a 50:50 mixture of native and exogenous lipids significantly altered the orientation of Na(+) K(+)-ATPase within the membrane.

Conclusions:

  • The lipid environment of sperm membranes critically influences the orientation of membrane proteins like Na(+) K(+)-ATPase.
  • Changes in lipid composition can lead to altered protein positioning, potentially impacting sperm function and fertilization capabilities.
  • This study provides insights into the molecular mechanisms underlying sperm capacitation and the role of membrane dynamics.