Watching conformational- and photodynamics of single fluorescent proteins in solution
Open Access
- 31 January 2010
- journal article
- research article
- Published by Springer Science and Business Media LLC in Nature Chemistry
- Vol. 2 (3), 179-186
- https://doi.org/10.1038/nchem.545
Abstract
Observing the dynamics of single biomolecules over prolonged time periods is difficult to achieve without significantly altering the molecule through immobilization. It can, however, be accomplished using the anti-Brownian electrokinetic trap, which allows extended investigation of solution-phase biomolecules—without immobilization—through real-time electrokinetic feedback. Here we apply the trap to study an important photosynthetic antenna protein, allophycocyanin. The technique allows the observation of single molecules of solution-phase allophycocyanin for more than one second. We observe a complex relationship between fluorescence intensity and lifetime that cannot be explained by simple static kinetic models. Light-induced conformational changes are shown to occur and evidence is obtained for fluctuations in the spontaneous emission lifetime, which is typically assumed to be constant. Our methods provide a new window into the dynamics of fluorescent proteins and the observations are relevant for the interpretation of in vivo single-molecule imaging experiments, bacterial photosynthetic regulation and biomaterials for solar energy harvesting.Keywords
This publication has 51 references indexed in Scilit:
- Photoinduced Unfolding of β-Lactoglobulin Mediated by a Water-Soluble PorphyrinThe Journal of Physical Chemistry B, 2009
- Activation of CaMKII in single dendritic spines during long-term potentiationNature, 2009
- Cytosolic Viral Sensor RIG-I Is a 5'-Triphosphate–Dependent Translocase on Double-Stranded RNAScience, 2009
- Visualization of Membrane Rafts Using a Perylene Monoimide Derivative and Fluorescence Lifetime ImagingBiophysical Journal, 2007
- Principal-components analysis of shape fluctuations of single DNA moleculesProceedings of the National Academy of Sciences of the United States of America, 2007
- Analysis of Single-Molecule FRET Trajectories Using Hidden Markov ModelingBiophysical Journal, 2006
- Effects of surface tethering on protein folding mechanismsProceedings of the National Academy of Sciences, 2006
- In all likelihood: Statistical modelling and inference using likelihood, by Yudi Pawitan. Pp. 528. £40. 2001. ISBN 0 19 850765 8 (Oxford University Press).The Mathematical Gazette, 2002
- Isolation, crystallization, crystal structure analysis and refinement of allophycocyanin from the cyanobacterium Spirulina platensis at 2.3 Å resolutionJournal of Molecular Biology, 1995
- Synthesis of bile pigments. 14. Synthesis of a bilindionostilbenoparacyclophane as a model for stretched bile pigment chromophores of biliproteinsJournal of the American Chemical Society, 1987