Mechanical Unfolding Pathways of the Enhanced Yellow Fluorescent Protein Revealed by Single Molecule Force Spectroscopy.
From: Department of Biological Sciences, Columbia University, New York, New York 10027, USA.
The Journal of biological chemistry
- Publish Date: Dec 2006
- ISSN: 0021-9258
- Volume: 281
- Issue: 52
- Pages: 40010-4
- Medium: Print
- Language: English
- Citation (JAMA): Perez-Jimenez Raul, Garcia-Manyes Sergi, Ainavarapu Sri Rama Koti, et al. Mechanical Unfolding Pathways of the Enhanced Yellow Fluorescent Protein Revealed by Single Molecule Force Spectroscopy.. J. Biol. Chem. Dec 2006;281:40010-4
Abstract
We used single molecule force spectroscopy to characterize the mechanical stability of the enhanced yellow fluorescent protein (EYFP) (a mutant form of the green fluorescent protein (GFP)) and two of its circularly permutated variants. In all three constructs, we found two main unfolding peaks; the first corresponds to a transition state placed close to the termini and the second to a transition state placed halfway through the molecule. We attribute the second transition state to the shear rupture of the beta1- and beta6-strands, which we verified by introducing a point mutation in this region. Although both unfolding peaks were observed in all three EYFP variants, their relative frequency of occurrence varied. Our results demonstrated that the mechanical unfolding pathways in EYFP could be deciphered through the use of circular permutation.
Mesh Headings (Keywords): Animals, Green Fluorescent Proteins, Microscopy, Atomic Force, Protein Engineering, Protein Folding, Scyphozoa, Signal Transduction
Check for Full Text / PubMed Unique Identifier (PMID): 17082195
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