2fwq
From Proteopedia
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| - | [[Image:2fwq.gif|left|200px]]<br /><applet load="2fwq" size="350" color="white" frame="true" align="right" spinBox="true" | ||
| - | caption="2fwq, resolution 1.40Å" /> | ||
| - | '''Reduced enolate chromophore intermediate for Y66H GFP variant'''<br /> | ||
| - | == | + | ==Reduced enolate chromophore intermediate for Y66H GFP variant== |
| + | <StructureSection load='2fwq' size='340' side='right'caption='[[2fwq]], [[Resolution|resolution]] 1.40Å' scene=''> | ||
| + | == Structural highlights == | ||
| + | <table><tr><td colspan='2'>[[2fwq]] is a 1 chain structure with sequence from [https://en.wikipedia.org/wiki/Aequorea_victoria Aequorea victoria]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=2FWQ OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=2FWQ FirstGlance]. <br> | ||
| + | </td></tr><tr id='method'><td class="sblockLbl"><b>[[Empirical_models|Method:]]</b></td><td class="sblockDat" id="methodDat">X-ray diffraction, [[Resolution|Resolution]] 1.4Å</td></tr> | ||
| + | <tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=MG:MAGNESIUM+ION'>MG</scene>, <scene name='pdbligand=XXY:2-[(1R,2R)-1-AMINO-2-HYDROXYPROPYL]-1-(CARBOXYMETHYL)-4-(1H-IMIDAZOL-5-YLMETHYL)-1H-IMIDAZOL-5-OLATE'>XXY</scene></td></tr> | ||
| + | <tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=2fwq FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=2fwq OCA], [https://pdbe.org/2fwq PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=2fwq RCSB], [https://www.ebi.ac.uk/pdbsum/2fwq PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=2fwq ProSAT]</span></td></tr> | ||
| + | </table> | ||
| + | == Function == | ||
| + | [https://www.uniprot.org/uniprot/GFP_AEQVI GFP_AEQVI] Energy-transfer acceptor. Its role is to transduce the blue chemiluminescence of the protein aequorin into green fluorescent light by energy transfer. Fluoresces in vivo upon receiving energy from the Ca(2+)-activated photoprotein aequorin. | ||
| + | == Evolutionary Conservation == | ||
| + | [[Image:Consurf_key_small.gif|200px|right]] | ||
| + | Check<jmol> | ||
| + | <jmolCheckbox> | ||
| + | <scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/fw/2fwq_consurf.spt"</scriptWhenChecked> | ||
| + | <scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview03.spt</scriptWhenUnchecked> | ||
| + | <text>to colour the structure by Evolutionary Conservation</text> | ||
| + | </jmolCheckbox> | ||
| + | </jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=2fwq ConSurf]. | ||
| + | <div style="clear:both"></div> | ||
| + | <div style="background-color:#fffaf0;"> | ||
| + | == Publication Abstract from PubMed == | ||
The Aequorea victoria green fluorescent protein (GFP) creates a fluorophore from its component amino acids Ser65, Tyr66, and Gly67 through a remarkable post-translational modification, involving spontaneous peptide backbone cyclization, dehydration, and oxidation reactions. Here we test and extend the understanding of fluorophore biosynthesis by coupling chemical reduction and anaerobic methodologies with kinetic analyses and protein structure determination. Two high-resolution structures of dithionite-treated GFP variants reveal a previously uncharacterized enolate intermediate form of the chromophore that is viable in generating a fluorophore (t1/2 = 39 min-1) upon exposure to air. Isolation of this enolate intermediate will now allow specific probing of the rate-limiting oxidation step for fluorophore biosynthesis in GFP and its red fluorescent protein homologues. Such targeted characterizations may lead to the design of faster maturing proteins with enhanced applications in biotechnology and cell biology. Moreover, our results reveal how the GFP protein environment mimics enzyme systems, by stabilizing an otherwise high energy enolate intermediate to achieve its post-translational modification. | The Aequorea victoria green fluorescent protein (GFP) creates a fluorophore from its component amino acids Ser65, Tyr66, and Gly67 through a remarkable post-translational modification, involving spontaneous peptide backbone cyclization, dehydration, and oxidation reactions. Here we test and extend the understanding of fluorophore biosynthesis by coupling chemical reduction and anaerobic methodologies with kinetic analyses and protein structure determination. Two high-resolution structures of dithionite-treated GFP variants reveal a previously uncharacterized enolate intermediate form of the chromophore that is viable in generating a fluorophore (t1/2 = 39 min-1) upon exposure to air. Isolation of this enolate intermediate will now allow specific probing of the rate-limiting oxidation step for fluorophore biosynthesis in GFP and its red fluorescent protein homologues. Such targeted characterizations may lead to the design of faster maturing proteins with enhanced applications in biotechnology and cell biology. Moreover, our results reveal how the GFP protein environment mimics enzyme systems, by stabilizing an otherwise high energy enolate intermediate to achieve its post-translational modification. | ||
| - | + | Structural evidence for an enolate intermediate in GFP fluorophore biosynthesis.,Barondeau DP, Tainer JA, Getzoff ED J Am Chem Soc. 2006 Mar 15;128(10):3166-8. PMID:16522096<ref>PMID:16522096</ref> | |
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| - | + | From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br> | |
| - | + | </div> | |
| - | + | <div class="pdbe-citations 2fwq" style="background-color:#fffaf0;"></div> | |
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| - | + | ==See Also== | |
| + | *[[Green Fluorescent Protein 3D structures|Green Fluorescent Protein 3D structures]] | ||
| + | == References == | ||
| + | <references/> | ||
| + | __TOC__ | ||
| + | </StructureSection> | ||
| + | [[Category: Aequorea victoria]] | ||
| + | [[Category: Large Structures]] | ||
| + | [[Category: Barondeau DP]] | ||
| + | [[Category: Getzoff ED]] | ||
| + | [[Category: Tainer JA]] | ||
Current revision
Reduced enolate chromophore intermediate for Y66H GFP variant
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