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| | ==Engineered Fab fragment specific for EYMPME (EE) peptide== | | ==Engineered Fab fragment specific for EYMPME (EE) peptide== |
| - | <StructureSection load='4x0k' size='340' side='right' caption='[[4x0k]], [[Resolution|resolution]] 2.04Å' scene=''> | + | <StructureSection load='4x0k' size='340' side='right'caption='[[4x0k]], [[Resolution|resolution]] 2.04Å' scene=''> |
| | == Structural highlights == | | == Structural highlights == |
| - | <table><tr><td colspan='2'>[[4x0k]] is a 4 chain structure with sequence from [http://en.wikipedia.org/wiki/Human Human]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=4X0K OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=4X0K FirstGlance]. <br> | + | <table><tr><td colspan='2'>[[4x0k]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Homo_sapiens Homo sapiens]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=4X0K OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=4X0K FirstGlance]. <br> |
| - | </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=4x0k FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=4x0k OCA], [http://pdbe.org/4x0k PDBe], [http://www.rcsb.org/pdb/explore.do?structureId=4x0k RCSB], [http://www.ebi.ac.uk/pdbsum/4x0k PDBsum], [http://prosat.h-its.org/prosat/prosatexe?pdbcode=4x0k ProSAT]</span></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]] 2.04Å</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=4x0k FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=4x0k OCA], [https://pdbe.org/4x0k PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=4x0k RCSB], [https://www.ebi.ac.uk/pdbsum/4x0k PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=4x0k ProSAT]</span></td></tr> |
| | </table> | | </table> |
| | <div style="background-color:#fffaf0;"> | | <div style="background-color:#fffaf0;"> |
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| | __TOC__ | | __TOC__ |
| | </StructureSection> | | </StructureSection> |
| - | [[Category: Human]] | + | [[Category: Homo sapiens]] |
| - | [[Category: Johnson, J L]] | + | [[Category: Large Structures]] |
| - | [[Category: Lieberman, R L]] | + | [[Category: Johnson JL]] |
| - | [[Category: Antibody fragment]] | + | [[Category: Lieberman RL]] |
| - | [[Category: Crystallization chaperone]]
| + | |
| - | [[Category: Fab fragment]]
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| - | [[Category: Immune system]]
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| Structural highlights
Publication Abstract from PubMed
Crystallization chaperones are attracting increasing interest as a route to crystal growth and structure elucidation of difficult targets such as membrane proteins. While strategies to date have typically employed protein-specific chaperones, a peptide-specific chaperone to crystallize multiple cognate peptide epitope-containing client proteins is envisioned. This would eliminate the target-specific chaperone-production step and streamline the co-crystallization process. Previously, protein engineering and directed evolution were used to generate a single-chain variable (scFv) antibody fragment with affinity for the peptide sequence EYMPME (scFv/EE). This report details the conversion of scFv/EE to an anti-EE Fab format (Fab/EE) followed by its biophysical characterization. The addition of constant chains increased the overall stability and had a negligible impact on the antigen affinity. The 2.0 A resolution crystal structure of Fab/EE reveals contacts with larger surface areas than those of scFv/EE. Surface plasmon resonance, an enzyme-linked immunosorbent assay, and size-exclusion chromatography were used to assess Fab/EE binding to EE-tagged soluble and membrane test proteins: namely, the beta-barrel outer membrane protein intimin and alpha-helical A2a G protein-coupled receptor (A2aR). Molecular-dynamics simulation of the intimin constructs with and without Fab/EE provides insight into the energetic complexities of the co-crystallization approach.
Structural and biophysical characterization of an epitope-specific engineered Fab fragment and complexation with membrane proteins: implications for co-crystallization.,Johnson JL, Entzminger KC, Hyun J, Kalyoncu S, Heaner DP Jr, Morales IA, Sheppard A, Gumbart JC, Maynard JA, Lieberman RL Acta Crystallogr D Biol Crystallogr. 2015 Apr;71(Pt 4):896-906. doi:, 10.1107/S1399004715001856. Epub 2015 Mar 27. PMID:25849400[1]
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.
References
- ↑ Johnson JL, Entzminger KC, Hyun J, Kalyoncu S, Heaner DP Jr, Morales IA, Sheppard A, Gumbart JC, Maynard JA, Lieberman RL. Structural and biophysical characterization of an epitope-specific engineered Fab fragment and complexation with membrane proteins: implications for co-crystallization. Acta Crystallogr D Biol Crystallogr. 2015 Apr;71(Pt 4):896-906. doi:, 10.1107/S1399004715001856. Epub 2015 Mar 27. PMID:25849400 doi:http://dx.doi.org/10.1107/S1399004715001856
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