This old version of Proteopedia is provided for student assignments while the new version is undergoing repairs. Content and edits done in this old version of Proteopedia after March 1, 2026 will eventually be lost when it is retired in about June of 2026.
Apply for new accounts at the new Proteopedia. Your logins will work in both the old and new versions.
1h6r
From Proteopedia
(New page: 200px<br /><applet load="1h6r" size="450" color="white" frame="true" align="right" spinBox="true" caption="1h6r, resolution 1.50Å" /> '''THE OXIDIZED STATE O...) |
|||
| Line 1: | Line 1: | ||
| - | [[Image:1h6r.gif|left|200px]]<br /><applet load="1h6r" size=" | + | [[Image:1h6r.gif|left|200px]]<br /><applet load="1h6r" size="350" color="white" frame="true" align="right" spinBox="true" |
caption="1h6r, resolution 1.50Å" /> | caption="1h6r, resolution 1.50Å" /> | ||
'''THE OXIDIZED STATE OF A REDOX SENSITIVE VARIANT OF GREEN FLUORESCENT PROTEIN'''<br /> | '''THE OXIDIZED STATE OF A REDOX SENSITIVE VARIANT OF GREEN FLUORESCENT PROTEIN'''<br /> | ||
==Overview== | ==Overview== | ||
| - | To visualize the formation of disulfide bonds in living cells, a pair of | + | To visualize the formation of disulfide bonds in living cells, a pair of redox-active cysteines was introduced into the yellow fluorescent variant of green fluorescent protein. Formation of a disulfide bond between the two cysteines was fully reversible and resulted in a >2-fold decrease in the intrinsic fluorescence. Inter conversion between the two redox states could thus be followed in vitro as well as in vivo by non-invasive fluorimetric measurements. The 1.5 A crystal structure of the oxidized protein revealed a disulfide bond-induced distortion of the beta-barrel, as well as a structural reorganization of residues in the immediate chromophore environment. By combining this information with spectroscopic data, we propose a detailed mechanism accounting for the observed redox state-dependent fluorescence. The redox potential of the cysteine couple was found to be within the physiological range for redox-active cysteines. In the cytoplasm of Escherichia coli, the protein was a sensitive probe for the redox changes that occur upon disruption of the thioredoxin reductive pathway. |
==About this Structure== | ==About this Structure== | ||
| - | 1H6R is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Aequorea_victoria Aequorea victoria] with CL as [http://en.wikipedia.org/wiki/ligand ligand]. Full crystallographic information is available from [http:// | + | 1H6R is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Aequorea_victoria Aequorea victoria] with <scene name='pdbligand=CL:'>CL</scene> as [http://en.wikipedia.org/wiki/ligand ligand]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1H6R OCA]. |
==Reference== | ==Reference== | ||
| Line 13: | Line 13: | ||
[[Category: Aequorea victoria]] | [[Category: Aequorea victoria]] | ||
[[Category: Single protein]] | [[Category: Single protein]] | ||
| - | [[Category: Hansen, F | + | [[Category: Hansen, F G.]] |
[[Category: Henriksen, A.]] | [[Category: Henriksen, A.]] | ||
[[Category: Ostergaard, H.]] | [[Category: Ostergaard, H.]] | ||
| - | [[Category: Winther, J | + | [[Category: Winther, J R.]] |
[[Category: CL]] | [[Category: CL]] | ||
[[Category: green fluorescent protein]] | [[Category: green fluorescent protein]] | ||
| Line 22: | Line 22: | ||
[[Category: yellow-emission]] | [[Category: yellow-emission]] | ||
| - | ''Page seeded by [http:// | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 12:58:00 2008'' |
Revision as of 10:58, 21 February 2008
|
THE OXIDIZED STATE OF A REDOX SENSITIVE VARIANT OF GREEN FLUORESCENT PROTEIN
Overview
To visualize the formation of disulfide bonds in living cells, a pair of redox-active cysteines was introduced into the yellow fluorescent variant of green fluorescent protein. Formation of a disulfide bond between the two cysteines was fully reversible and resulted in a >2-fold decrease in the intrinsic fluorescence. Inter conversion between the two redox states could thus be followed in vitro as well as in vivo by non-invasive fluorimetric measurements. The 1.5 A crystal structure of the oxidized protein revealed a disulfide bond-induced distortion of the beta-barrel, as well as a structural reorganization of residues in the immediate chromophore environment. By combining this information with spectroscopic data, we propose a detailed mechanism accounting for the observed redox state-dependent fluorescence. The redox potential of the cysteine couple was found to be within the physiological range for redox-active cysteines. In the cytoplasm of Escherichia coli, the protein was a sensitive probe for the redox changes that occur upon disruption of the thioredoxin reductive pathway.
About this Structure
1H6R is a Single protein structure of sequence from Aequorea victoria with as ligand. Full crystallographic information is available from OCA.
Reference
Shedding light on disulfide bond formation: engineering a redox switch in green fluorescent protein., Ostergaard H, Henriksen A, Hansen FG, Winther JR, EMBO J. 2001 Nov 1;20(21):5853-62. PMID:11689426
Page seeded by OCA on Thu Feb 21 12:58:00 2008
