Protein denaturation at the air-water interface and how to prevent it

Elife. 2019 Apr 1:8:e42747. doi: 10.7554/eLife.42747.

Abstract

Electron cryo-microscopy analyzes the structure of proteins and protein complexes in vitrified solution. Proteins tend to adsorb to the air-water interface in unsupported films of aqueous solution, which can result in partial or complete denaturation. We investigated the structure of yeast fatty acid synthase at the air-water interface by electron cryo-tomography and single-particle image processing. Around 90% of complexes adsorbed to the air-water interface are partly denatured. We show that the unfolded regions face the air-water interface. Denaturation by contact with air may happen at any stage of specimen preparation. Denaturation at the air-water interface is completely avoided when the complex is plunge-frozen on a substrate of hydrophilized graphene.

Keywords: CryoEM; CryoET; Graphene; Protein denaturation; S. cerevisiae; molecular biophysics; structural biology.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adsorption
  • Air*
  • Cryoelectron Microscopy
  • Electron Microscope Tomography
  • Fatty Acid Synthases / chemistry*
  • Protein Denaturation*
  • Saccharomyces cerevisiae Proteins / chemistry*
  • Water / chemistry*

Substances

  • Saccharomyces cerevisiae Proteins
  • Water
  • Fatty Acid Synthases