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Related Experiment Videos

Brominated detergents as tools to study protein-detergent interactions

B De Foresta1, N Legros, D Plusquellec

  • 1Département de Biologie Cellulaire et Moléculaire (Commissariat à l'Energie Atomique, Centre d'Etudes de Saclay, Gif-sur-Yvette, France.

European Journal of Biochemistry
|October 15, 1996
PubMed
Summary

Brominated detergents effectively study protein-detergent interactions by quenching reticulum sarcoplasmic (SR) Ca(2+)-ATPase fluorescence. These novel detergents offer insights into hydrophobic surface contacts in membrane proteins.

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Area of Science:

  • Biochemistry
  • Biophysics
  • Membrane Protein Chemistry

Background:

  • Protein-detergent interactions are crucial for understanding membrane protein structure and function.
  • Non-ionic detergents are commonly used to solubilize membrane proteins, but their interactions can be complex.
  • Studying these interactions requires specific tools to probe the protein-detergent interface.

Purpose of the Study:

  • To synthesize and characterize brominated analogs of non-ionic detergents.
  • To investigate the short-range interactions between these brominated detergents and sarcoplasmic reticulum (SR) Ca(2+)-ATPase.
  • To evaluate the utility of brominated detergents as probes for protein-detergent interactions.

Main Methods:

  • Synthesis of 7,8-dibromododecyl beta-maltoside and 2-O-(10,11-dibromoundecanoyl)sucrose.

Related Experiment Videos

  • Rayleigh scattering measurements to assess solubilization efficiency and critical micellar concentration.
  • Fluorescence quenching assays using intrinsic fluorescence of SR Ca(2+)-ATPase.
  • Measurement of partition coefficients (K) between lipid and aqueous phases.
  • Enzyme activity assays (hydrolysis of p-nitrophenyl phosphate).

    • Main Results:

    • Brominated detergents efficiently solubilized SR vesicles with slightly higher critical micellar concentrations than their non-brominated counterparts.
    • Partition coefficients were comparable for analogs and parent detergents, indicating similar membrane partitioning.
    • Fluorescence quenching of Ca(2+)-ATPase increased with brominated detergent concentration, revealing significant protein-detergent contacts.
    • These contacts persisted through membrane solubilization and delipidation, with reversible quenching upon addition of non-brominated detergent.
    • Detergent effects on Ca(2+)-ATPase activity correlated with observed protein-detergent contacts.

    Conclusions:

    • Brominated detergents are effective tools for studying protein-detergent interactions at the hydrophobic surface of membrane proteins.
    • These detergents provide valuable insights into interactions both within membranes and in solubilized complexes.
    • The fluorescence quenching method offers a sensitive way to quantify protein-detergent contacts.