Staff Publications

  • external user (warningwarning)
  • Log in as
  • language uk
Record number 534474
Title Maximum entropy analysis of polarized fluorescence decay of (E)GFP in aqueous solution
Author(s) Novikov, Eugene G.; Skakun, Victor V.; Borst, Jan Willem; Visser, Antonie J.W.G.
Source Methods and Applications in Fluorescence 6 (2018)1. - ISSN 2050-6120
DOI http://dx.doi.org/10.1088/2050-6120/aa898b
Department(s) Biochemistry
VLAG
EPS
Publication type Refereed Article in a scientific journal
Publication year 2018
Keyword(s) (enhanced) green fluorescent protein - Maximum entropy method - Rate spectrum - Rotational correlation time - Rotational diffusion - Time-resolved fluorescence - Time-resolved fluorescence anisotropy
Abstract The maximum entropy method (MEM) was used for the analysis of polarized fluorescence decays of enhanced green fluorescent protein (EGFP) in buffered water/glycerol mixtures, obtained with time-correlated single-photon counting (Visser et al 2016 Methods Appl. Fluoresc. 4 035002). To this end, we used a general-purpose software module of MEM that was earlier developed to analyze (complex) laser photolysis kinetics of ligand rebinding reactions in oxygen binding proteins. We demonstrate that the MEM software provides reliable results and is easy to use for the analysis of both total fluorescence decay and fluorescence anisotropy decay of aqueous solutions of EGFP. The rotational correlation times of EGFP in water/glycerol mixtures, obtained by MEM as maxima of the correlation-time distributions, are identical to the single correlation times determined by global analysis of parallel and perpendicular polarized decay components. The MEM software is also able to determine homo-FRET in another dimeric GFP, for which the transfer correlation time is an order of magnitude shorter than the rotational correlation time. One important advantage utilizing MEM analysis is that no initial guesses of parameters are required, since MEM is able to select the least correlated solution from the feasible set of solutions.
Comments
There are no comments yet. You can post the first one!
Post a comment
 
Please log in to use this service. Login as Wageningen University & Research user or guest user in upper right hand corner of this page.