publication . Article . 2012

Fluorescence Fluctuation Spectroscopy Applied to Cell-Free Expression, Chromophore Maturation and Protein-DNA Interaction

Macdonald, Patrick J.; Chen, Yan; Li, Ming; Harris, Reuben S.; Mueller, Joachim D.;
Open Access
  • Published: 01 Jan 2012 Journal: Biophysical Journal, volume 102, issue 3 (issn: 0006-3495, Copyright policy)
  • Publisher: Elsevier BV
Abstract
Cell-free expression systems are increasingly being employed as platforms for biophysical, biochemical, and systems biology experiments. We demonstrate that combining fluorescence fluctuation spectroscopy (FFS) with cell-free expression provides quick and quantitative measurements of chromophore maturation, protein-protein interactions and protein-DNA interactions. We perform the first study of chromophore maturation as a function of temperature, and demonstrate pronounced temperature dependence of the maturation kinetics for EGFP, EYFP, and mCherry. The Eyring equation successfully reproduces the temperature-dependence of the maturation rate for each of the pro...
Subjects
free text keywords: Biophysics, Protein–DNA interaction, Fluorescence, DNA, chemistry.chemical_compound, chemistry, mCherry, Kinetics, Chromophore, Green fluorescent protein, A-DNA, Biochemistry
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