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...) |
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| - | [[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== | ||
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[[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]] | ||
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[[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
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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
