Probing the self-assembly and the accompanying structural changes of hydrophobin SC3 on a hydrophobic surface by mass spectrometry

Biophys J. 2004 Sep;87(3):1919-28. doi: 10.1529/biophysj.104.041616.

Abstract

The fungal class I hydrophobin SC3 self-assembles into an amphipathic membrane at hydrophilic-hydrophobic interfaces such as the water-air and water-Teflon interface. During self-assembly, the water-soluble state of SC3 proceeds via the intermediate alpha-helical state to the stable end form called the beta-sheet state. Self-assembly of the hydrophobin at the Teflon surface is arrested in the alpha-helical state. The beta-sheet state can be induced at elevated temperature in the presence of detergent. The structural changes of SC3 were monitored by various mass spectrometry techniques. We show that the so-called second loop of SC3 (C39-S72) has a high affinity for Teflon. Binding of this part of SC3 to Teflon was accompanied by the formation of alpha-helical structure and resulted in low solvent accessibility. The solvent-protected region of the second loop extended upon conversion to the beta-sheet state. In contrast, the C-terminal part of SC3 became more exposed to the solvent. The results indicate that the second loop of class I hydrophobins plays a pivotal role in self-assembly at the hydrophilic-hydrophobic interface. Of interest, this loop is much smaller in case of class II hydrophobins, which may explain the differences in their assembly.

MeSH terms

  • Air
  • Amino Acid Sequence
  • Circular Dichroism
  • Detergents / pharmacology
  • Endopeptidases / pharmacology
  • Formates / chemistry
  • Fungal Proteins / chemistry*
  • Kinetics
  • Mass Spectrometry / methods*
  • Metalloendopeptidases
  • Molecular Sequence Data
  • Oxygen / metabolism
  • Pepsin A / pharmacology
  • Peptides / chemistry
  • Polytetrafluoroethylene / chemistry
  • Protein Binding
  • Protein Conformation
  • Protein Structure, Secondary
  • Protein Structure, Tertiary
  • Sequence Homology, Amino Acid
  • Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
  • Temperature
  • Time Factors
  • Water

Substances

  • Detergents
  • Formates
  • Fungal Proteins
  • Peptides
  • Water
  • peroxyformic acid
  • Polytetrafluoroethylene
  • Endopeptidases
  • Pepsin A
  • Metalloendopeptidases
  • endoproteinase Asp-N
  • Oxygen