Tracking unfolding and refolding reactions of single proteins using atomic force microscopy methods

Research output: Contribution to journalArticle

14 Citations (Scopus)

Abstract

During the last two decades single-molecule manipulation techniques such as atomic force microscopy (AFM) has risen to prominence through their unique capacity to provide fundamental information on the structure and function of biomolecules. Here we describe the use of single-molecule AFM to track protein unfolding and refolding pathways, enzymatic catalysis and the effects of osmolytes and chaperones on protein stability and folding. We will outline the principles of operation for two different AFM pulling techniques: length clamp and force-clamp and discuss prominent applications. We provide protocols for the construction of polyproteins which are amenable for AFM experiments, the preparation of different coverslips, choice and calibration of AFM cantilevers. We also discuss the selection criteria for AFM recordings, the calibration of AFM cantilevers, protein sample preparations and analysis of the obtained data.

Original languageEnglish (US)
Pages (from-to)151-160
Number of pages10
JournalMethods
Volume60
Issue number2
DOIs
StatePublished - Apr 1 2013

Fingerprint

Atomic Force Microscopy
Atomic force microscopy
Proteins
Clamping devices
Calibration
Protein Refolding
Polyproteins
Protein Unfolding
Molecules
Protein Stability
Protein Folding
Biomolecules
Catalysis
Patient Selection

Keywords

  • Atomic force microscopy
  • Biophysics
  • Chaperones
  • Osmolytes
  • Protein folding
  • Single-molecule
  • Unfolding

ASJC Scopus subject areas

  • Molecular Biology
  • Biochemistry, Genetics and Molecular Biology(all)

Cite this

Tracking unfolding and refolding reactions of single proteins using atomic force microscopy methods. / Bujalowski, Pawel; Oberhauser, Andres.

In: Methods, Vol. 60, No. 2, 01.04.2013, p. 151-160.

Research output: Contribution to journalArticle

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