1l7i
From Proteopedia
(New page: 200px<br /> <applet load="1l7i" size="450" color="white" frame="true" align="right" spinBox="true" caption="1l7i, resolution 1.80Å" /> '''Crystal Structure o...) |
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| - | [[Image:1l7i.gif|left|200px]]<br /> | + | [[Image:1l7i.gif|left|200px]]<br /><applet load="1l7i" size="350" color="white" frame="true" align="right" spinBox="true" |
| - | <applet load="1l7i" size=" | + | |
caption="1l7i, resolution 1.80Å" /> | caption="1l7i, resolution 1.80Å" /> | ||
'''Crystal Structure of the anti-ErbB2 Fab2C4'''<br /> | '''Crystal Structure of the anti-ErbB2 Fab2C4'''<br /> | ||
==Overview== | ==Overview== | ||
| - | Shotgun scanning combinatorial mutagenesis was used to study the | + | Shotgun scanning combinatorial mutagenesis was used to study the antigen-binding site of Fab2C4, a humanized monoclonal antibody fragment that binds to the extracellular domain of the human oncogene product ErbB2. Essentially all the residues in the Fab2C4 complementarity determining regions (CDRs) were alanine-scanned using phage-displayed libraries that preferentially allowed side-chains to vary as the wild-type or alanine. A separate homolog-scan was performed using libraries that allowed side-chains to vary only as the wild-type or a similar amino acid residue. Following binding selections to isolate functional clones, DNA sequencing was used to determine the wild-type/mutant ratios at each varied position, and these ratios were used to assess the contributions of each side-chain to antigen binding. The alanine-scan revealed that most of the side-chains that contribute to antigen binding are located in the heavy chain, and the Fab2C4 three-dimensional structure revealed that these residues fall into two groups. The first group consists of solvent-exposed residues which likely make energetically favorable contacts with the antigen and thus comprise the functional-binding epitope. The second group consists of buried residues with side-chains that pack against other CDR residues and apparently act as scaffolding to maintain the functional epitope in a binding-competent conformation. The homolog-scan involved subtle mutations, and as a result, only a subset of the side-chains that were intolerant to alanine substitutions were also intolerant to homologous substitutions. In particular, the 610 A2 functional epitope surface revealed by alanine-scanning shrunk to only 369 A2 when mapped with homologous substitutions, suggesting that this smaller subset of side-chains may be involved in more precise contacts with the antigen. The results validate shotgun scanning as a rapid and accurate method for determining the functional contributions of individual side-chains involved in protein-protein interactions. |
==About this Structure== | ==About this Structure== | ||
| - | 1L7I is a [http://en.wikipedia.org/wiki/Protein_complex Protein complex] structure of sequences from [http://en.wikipedia.org/wiki/Homo_sapiens Homo sapiens] with SO4 as [http://en.wikipedia.org/wiki/ligand ligand]. Full crystallographic information is available from [http:// | + | 1L7I is a [http://en.wikipedia.org/wiki/Protein_complex Protein complex] structure of sequences from [http://en.wikipedia.org/wiki/Homo_sapiens Homo sapiens] with <scene name='pdbligand=SO4:'>SO4</scene> as [http://en.wikipedia.org/wiki/ligand ligand]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1L7I OCA]. |
==Reference== | ==Reference== | ||
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[[Category: Homo sapiens]] | [[Category: Homo sapiens]] | ||
[[Category: Protein complex]] | [[Category: Protein complex]] | ||
| - | [[Category: Adams, C | + | [[Category: Adams, C W.]] |
| - | [[Category: Breece, T | + | [[Category: Breece, T N.]] |
| - | [[Category: Presta, L | + | [[Category: Presta, L G.]] |
| - | [[Category: Sidhu, S | + | [[Category: Sidhu, S S.]] |
| - | [[Category: Vajdos, F | + | [[Category: Vajdos, F F.]] |
| - | [[Category: Vos, A | + | [[Category: Vos, A M.de.]] |
[[Category: SO4]] | [[Category: SO4]] | ||
[[Category: fab fragment]] | [[Category: fab fragment]] | ||
[[Category: ig domain]] | [[Category: ig domain]] | ||
| - | ''Page seeded by [http:// | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 13:42:10 2008'' |
Revision as of 11:42, 21 February 2008
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Crystal Structure of the anti-ErbB2 Fab2C4
Overview
Shotgun scanning combinatorial mutagenesis was used to study the antigen-binding site of Fab2C4, a humanized monoclonal antibody fragment that binds to the extracellular domain of the human oncogene product ErbB2. Essentially all the residues in the Fab2C4 complementarity determining regions (CDRs) were alanine-scanned using phage-displayed libraries that preferentially allowed side-chains to vary as the wild-type or alanine. A separate homolog-scan was performed using libraries that allowed side-chains to vary only as the wild-type or a similar amino acid residue. Following binding selections to isolate functional clones, DNA sequencing was used to determine the wild-type/mutant ratios at each varied position, and these ratios were used to assess the contributions of each side-chain to antigen binding. The alanine-scan revealed that most of the side-chains that contribute to antigen binding are located in the heavy chain, and the Fab2C4 three-dimensional structure revealed that these residues fall into two groups. The first group consists of solvent-exposed residues which likely make energetically favorable contacts with the antigen and thus comprise the functional-binding epitope. The second group consists of buried residues with side-chains that pack against other CDR residues and apparently act as scaffolding to maintain the functional epitope in a binding-competent conformation. The homolog-scan involved subtle mutations, and as a result, only a subset of the side-chains that were intolerant to alanine substitutions were also intolerant to homologous substitutions. In particular, the 610 A2 functional epitope surface revealed by alanine-scanning shrunk to only 369 A2 when mapped with homologous substitutions, suggesting that this smaller subset of side-chains may be involved in more precise contacts with the antigen. The results validate shotgun scanning as a rapid and accurate method for determining the functional contributions of individual side-chains involved in protein-protein interactions.
About this Structure
1L7I is a Protein complex structure of sequences from Homo sapiens with as ligand. Full crystallographic information is available from OCA.
Reference
Comprehensive functional maps of the antigen-binding site of an anti-ErbB2 antibody obtained with shotgun scanning mutagenesis., Vajdos FF, Adams CW, Breece TN, Presta LG, de Vos AM, Sidhu SS, J Mol Biol. 2002 Jul 5;320(2):415-28. PMID:12079396
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