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==Loop grafting onto a highly stable FN3 scaffold==
==Loop grafting onto a highly stable FN3 scaffold==
<StructureSection load='5j7k' size='340' side='right' caption='[[5j7k]], [[Resolution|resolution]] 2.46&Aring;' scene=''>
<StructureSection load='5j7k' size='340' side='right'caption='[[5j7k]], [[Resolution|resolution]] 2.46&Aring;' scene=''>
== Structural highlights ==
== Structural highlights ==
<table><tr><td colspan='2'>[[5j7k]] is a 8 chain structure. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=5J7K OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=5J7K FirstGlance]. <br>
<table><tr><td colspan='2'>[[5j7k]] is a 8 chain structure with sequence from [https://en.wikipedia.org/wiki/Synthetic_construct Synthetic construct]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=5J7K OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=5J7K FirstGlance]. <br>
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat"><scene name='pdbligand=ZN:ZINC+ION'>ZN</scene></td></tr>
</td></tr><tr id='method'><td class="sblockLbl"><b>[[Empirical_models|Method:]]</b></td><td class="sblockDat" id="methodDat">X-ray diffraction, [[Resolution|Resolution]] 2.46&#8491;</td></tr>
<tr id='related'><td class="sblockLbl"><b>[[Related_structure|Related:]]</b></td><td class="sblockDat">[[5j7c|5j7c]]</td></tr>
<tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=ZN:ZINC+ION'>ZN</scene></td></tr>
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=5j7k FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=5j7k OCA], [http://pdbe.org/5j7k PDBe], [http://www.rcsb.org/pdb/explore.do?structureId=5j7k RCSB], [http://www.ebi.ac.uk/pdbsum/5j7k PDBsum], [http://prosat.h-its.org/prosat/prosatexe?pdbcode=5j7k ProSAT]</span></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=5j7k FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=5j7k OCA], [https://pdbe.org/5j7k PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=5j7k RCSB], [https://www.ebi.ac.uk/pdbsum/5j7k PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=5j7k ProSAT]</span></td></tr>
</table>
</table>
<div style="background-color:#fffaf0;">
== Publication Abstract from PubMed ==
The favorable biophysical attributes of non-antibody scaffolds make them attractive alternatives to monoclonal antibodies. However, due to the well-known stability-function trade-off, these gains tend to be marginal after functional selection. A notable example is the fibronectin Type III (FN3) domain, FNfn10, which has been previously evolved to bind lysozyme with 1 pM affinity (FNfn10-alpha-lys), but suffers from poor thermodynamic and kinetic stability. To explore this stability-function compromise further, we grafted the lysozyme-binding loops from FNfn10-alpha-lys onto our previously engineered, ultra-stable FN3 scaffold, FN3con The resulting variant (FN3con-alpha-lys) bound lysozyme with a markedly reduced affinity, but retained high levels of thermal stability. The crystal structure of FNfn10-alpha-lys in complex with lysozyme revealed unanticipated interactions at the protein-protein interface involving framework residues of FNfn10-alpha-lys, thus explaining the failure to transfer binding via loop grafting. Utilizing this structural information, we redesigned FN3con-alpha-lys and restored picomolar binding affinity to lysozyme, while maintaining thermodynamic stability (with a thermal melting temperature 2-fold higher than that of FNfn10-alpha-lys). FN3con therefore provides an exceptional window of stability to tolerate deleterious mutations, resulting in a substantial advantage for functional design. This study emphasizes the utility of consensus design for the generation of highly stable scaffolds for downstream protein engineering studies.
Circumventing the stability-function trade-off in an engineered FN3 domain.,Porebski BT, Conroy PJ, Drinkwater N, Schofield P, Vazquez-Lombardi R, Hunter MR, Hoke DE, Christ D, McGowan S, Buckle AM Protein Eng Des Sel. 2016 Aug 29. PMID:27578887<ref>PMID:27578887</ref>
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br>
</div>
<div class="pdbe-citations 5j7k" style="background-color:#fffaf0;"></div>
== References ==
<references/>
__TOC__
__TOC__
</StructureSection>
</StructureSection>
[[Category: Buckle, A M]]
[[Category: Large Structures]]
[[Category: Drinkwater, N]]
[[Category: Synthetic construct]]
[[Category: McGowan, S]]
[[Category: Buckle AM]]
[[Category: Porebski, B T]]
[[Category: Drinkwater N]]
[[Category: Fibronectin type iii]]
[[Category: McGowan S]]
[[Category: Fn3]]
[[Category: Porebski BT]]
[[Category: Loop grafting]]
[[Category: Protein binding]]
[[Category: Protein binding-hydrolase complex]]
[[Category: Rational design]]
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