We investigated the donor effects (mCerulean3 versus mTurquoise2.1) on the spectroscopy and dynamics of mCerulean3-linker-mCitrine constructs using integrated fluorescence spectroscopy methods. Here, mCerulean3 (a cyan fluorescent protein) and mCitrine (a yellow fluorescent protein) act as Förster resonance energy transfer (FRET) pair, separated by flexible linker region. We hypothesize that the construct with mTurquoise2.1 would have many advantages as a donor, which include a higher FRET efficiency as compared with the mCerulean3 due to the enhanced spectral overlap with mCitrine. To test this hypothesis, we used steady-state spectroscopy, time-resolved fluorescence, and fluorescence correlation spectroscopy of both mCerulean3-linker-mCitrine and mTurquoise2.1-linker-mCitrine to investigate the donor effect on the FRET efficiency and translational diffusion as a means for developing a rational design for hetero-FRET constructs for environmental sensing.
|Original language||English (US)|
|Title of host publication||Ultrafast Nonlinear Imaging and Spectroscopy VIII|
|Editors||Zhiwen Liu, Demetri Psaltis, Kebin Shi|
|State||Published - 2020|
|Event||Ultrafast Nonlinear Imaging and Spectroscopy VIII 2020 - Virtual, Online, United States|
Duration: Aug 24 2020 → Sep 4 2020
|Name||Proceedings of SPIE - The International Society for Optical Engineering|
|Conference||Ultrafast Nonlinear Imaging and Spectroscopy VIII 2020|
|Period||8/24/20 → 9/4/20|
Bibliographical noteFunding Information:
We would like to thank Robert Miller and Christin Libal for their technical help and useful discussion during the course of this project. E.D.S. and A.A.H. acknowledge the financial support provided by the University of Minnesota Grant-in-Aid, a Chancellor’s Small Grant, the Department of Chemistry and Biochemistry, the Swenson College of Science and Engineering, University of Minnesota Duluth. A.J.B. acknowledges the financial support of the Netherlands Organization for Scientific Research Vidi grant. C.P.A. was supported by teaching fellowships from the Department of Chemistry and Biochemistry, University of Minnesota Duluth. T.M.K. acknowledges the support of Mylan Radulovich Graduate Fellowship as well as the teaching fellowship from the Department of Physics and Astronomy, University of Minnesota Duluth. We further acknowledge the support from the Minnesota Supercomputing Institute (MSI) at the University of Minnesota.
© 2020 SPIE.
- Fluorescence correlation spectroscopy
- Time-resolved fluorescence