roGFP

from Wikipedia, the free encyclopedia
GFP from Aequorea victoria , wild-type form

roGFP ( English Redox-sensitive GFP ) is a modified variant of the green fluorescent protein , in which the fluorescence changes depending on the redox .

properties

In comparison to the wild-type form of GFP, roGFP has two cysteines in the β-barrel structure of GFP at positions 147 and 204 on two adjacent β-strands in the cytosolic variant of roGFP1 (mutations C48S, S147C). By oxidation of the thiol groups is of the cysteines cystine formed, which affects in a blue shift in fluorescence of the rogfp.

Applications

The roGFP is used, among other things, as a redox-dependent biosensor and as a reporter protein. With roGFP the redox potential and the amount of glutathione in the cytosol can be determined in vivo , indirectly also hydrogen peroxide . As a fusion protein of the roGFP with glutaredoxin 1, the fluorescence can be increased.

In the endoplasmic reticulum , the formation of ROS can be followed with roGFP2 (mutations C48S, S147C, S65T) .

Alternative optical detection methods use e.g. B. Resazurin (synonym AlamarBlue), dichlorofluorescein (DCF) or the proteins HyPer or rxYFP .

Web links

Individual evidence

  1. ↑ Hanson GT, Aggeler R, Oglesbee D, Cannon M, Capaldi RA, Tsien RY, Remington SJ: Investigating mitochondrial redox potential with redox-sensitive green fluorescent protein indicators . In: J Biol Chem . 279, No. 13, 2004, pp. 13044-53. doi : 10.1074 / jbc.M312846200 . PMID 14722062 .
  2. ↑ KA Lukyanov, VV Belousov: Genetically encoded fluorescent redox sensors. In: Biochimica et Biophysica Acta . Volume 1840, Number 2, February 2014, pp. 745-756, ISSN  0006-3002 . doi : 10.1016 / j.bbagen.2013.05.030 (free full text). PMID 23726987 .
  3. ↑ Schwarzlander M., Fricker, MD, Muller, C., Marty, L., Brach, T., Novak, J., Sweetlove, LJ, Hell, R., and Meyer, AJ: Confocal imaging of glutathione redox potential in living plant cells . In: J Microsc . 279, No. 2, 2008, pp. 299-316. doi : 10.1111 / j.1365-2818.2008.02030.x . PMID 18778428 .
  4. ↑ CT Dooley, TM Dore, GT Hanson, WC Jackson, SJ Remington, RY Tsien: Imaging dynamic redox changes in mammalian cells with green fluorescent protein indicators. In: The Journal of biological chemistry. Volume 279, Number 21, May 2004, pp. 22284-22293, ISSN  0021-9258 . doi : 10.1074 / jbc.M312847200 . PMID 14985369 .
  5. ↑ Meyer, AJ, Brach, T., Marty, L., Kreye, S., Rouhier, N., Jacquot, JP, and Hell, R .: Redox-sensitive GFP in Arabidopsis thaliana is a quantitative biosensor for the redox potential of the cellular glutathione redox buffer . In: Plant J . 52, No. 5, 2007, pp. 973-86. doi : 10.1111 / j.1365-313X.2007.03280.x . PMID 17892447 .
  6. ^ SG Rhee, TS Chang, W. Jeong, D. Kang: Methods for detection and measurement of hydrogen peroxide inside and outside of cells. In: Molecules and cells. Volume 29, Number 6, June 2010, pp. 539-549, ISSN  0219-1032 . doi : 10.1007 / s10059-010-0082-3 . PMID 20526816 .
  7. ↑ Gutscher, M., Pauleau, AL, Marty, L., Brach, T., Wabnitz, GH, Saturday, Y., Meyer, AJ, and Dick, TP: Real-time imaging of the intracellular glutathione redox potential . In: Nat Methods . 5, No. 6, 2008, pp. 553-559. doi : 10.1038 / NMETH.1212 . PMID 18469822 .
  8. ↑ Brach T, Soyk S, Müller C, Hinz G, Hell R, Brandizzi F, Meyer AJ: Non-invasive topology analysis of membrane proteins in the secretory pathway . In: Plant J . 57, No. 3, 2009, pp. 534-41. doi : 10.1111 / j.1365-313X.2008.03704.x . PMID 18939964 .
  9. ↑ Schwarzländer M, Fricker MD, Sweetlove LJ: Monitoring the in vivo redox state of plant mitochondria: Effect of respiratory inhibitors, abiotic stress and assessment of recovery from oxidative challenge . In: Biochim Biophys Acta . 1787, No. 5, 2009, pp. 468-75. doi : 10.1016 / j.bbabio.2009.01.020 . PMID 19366606 .
  10. ↑ S. Pouvreau: Genetically encoded reactive oxygen species (ROS) and redox indicators. In: Biotechnology journal. Volume 9, Number 2, February 2014, pp. 282-293, ISSN  1860-7314 . doi : 10.1002 / biot.201300199 . PMID 24497389 .