Isolation of a Highly Reactive beta-Sheet-Rich Intermediate of Lysozyme in a Solvent-Free Liquid Phase

Alex P. S. Brogan, Kamendra P. Sharma, Adam W. Perriman*, Stephen Mann

*Corresponding author for this work

Research output: Contribution to journalArticle (Academic Journal)peer-review

20 Citations (Scopus)

Abstract

The thermal denaturation of solvent-free liquid lysozyme at temperatures in excess of 200 degrees C was studied by synchrotron radiation circular dichroism spectroscopy. Temperature-dependent changes in the secondary structure were used to map the equilibrium denaturation pathway and characterize a reactive beta-sheet-rich unfolding intermediate that was stable in the solvent-free liquid phase under anhydrous conditions but which underwent irreversible aggregation in the presence of water. The unfolding intermediate had a transition temperature of 78 degrees C and was extremely stable to temperature, eventually reaching the fully denatured state at 178 degrees C. We propose that the three-stage denaturation pathway arises from the decreased stability of the native state due to the absence of any appreciable hydrophobic effect, along with an entropically derived stabilization of the reactive intermediate associated with molecular crowding in the solvent-free liquid.

Original languageEnglish
Pages (from-to)8400-8407
Number of pages8
JournalJournal of Physical Chemistry B
Volume117
Issue number28
DOIs
Publication statusPublished - 18 Jul 2013

Keywords

  • CIRCULAR-DICHROISM SPECTROSCOPY
  • PROTEIN SECONDARY STRUCTURE
  • AMYLOID FIBRIL FORMATION
  • EGG-WHITE LYSOZYME
  • ORGANIC-SOLVENTS
  • STABILITY PARAMETERS
  • THERMAL-STABILITY
  • IONIC LIQUIDS
  • BIOCATALYSIS
  • CONFORMATION

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