Redesigning the Folding Pathway of a Model Three-helix Bundle Protein by Site-directed Mutagenesis.

Research output: Contribution to journalArticle


Because of their limited size and complexity, de novo designed proteins are particularly useful for the detailed investigation of folding thermodynamics and mechanisms. Here, we describe how subtle changes in the hydrophobic core of a model three-helix bundle protein (GM-0) alter its folding energetics. To explore the folding tolerance of GM-0 toward amino acid sequence variability, two mutant proteins (GM-1 and GM-2) were generated. In the mutants, cavities were created in the hydrophobic core of the protein by either singly (GM-1; L35A variant) or doubly (GM-2; L35A/I39A variant) replacing large hydrophobic side chains by smaller Ala residues. The folding of GM-0 is characterized by two partially folded intermediate states exhibiting characteristics of molten globules, as evidenced by pressure-unfolding and pressure-assisted cold denaturation experiments. In contrast, the folding energetics of both mutants, GM-1 and GM-2, exhibit only one folding intermediate. Our results support the view that simple but biologically important folding motifs such as the three-helix bundle can reveal complex folding plasticity, and they point to the role of hydrophobic packing as a determinant of the overall stability and folding thermodynamic of the helix bundle.


  • Dahabada H.J. Lopes
  • Alex Chapeaurouge
  • Gavin Manderson
  • Jonas S. Johansson
  • Sergio T. Ferreira
External organisations
  • University of Pennsylvania
Research areas and keywords

Subject classification (UKÄ) – MANDATORY

  • Rheumatology and Autoimmunity
Original languageEnglish
Pages (from-to)10991-10996
JournalJournal of Biological Chemistry
Issue number12
Publication statusPublished - 2004
Publication categoryResearch
Externally publishedYes

Bibliographic note

The information about affiliations in this record was updated in December 2015. The record was previously connected to the following departments: Connective Tissue Biology (013230151)