3b9v: Difference between revisions
New page: left|200px<br /><applet load="3b9v" size="350" color="white" frame="true" align="right" spinBox="true" caption="3b9v, resolution 1.800Å" /> '''Crystal Structure o... |
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'''Crystal Structure of an Autonomous VH Domain'''<br /> | '''Crystal Structure of an Autonomous VH Domain'''<br /> | ||
==Overview== | |||
We report a comprehensive analysis of sequence features that allow for the production of autonomous human heavy chain variable (V(H)) domains that are stable and soluble in the absence of a light chain partner. Using combinatorial phage-displayed libraries and conventional biophysical methods, we analyzed the entire former light chain interface and the third complementarity determining region (CDR3). Unlike the monomeric variable domains of camelid heavy chain antibodies (V(H)H domains), in which autonomous behavior depends on interactions between the hydrophobic former light chain interface and CDR3, we find that the stability of many in vitro evolved V(H) domains is essentially independent of the CDR3 sequence and instead derives from mutations that increase the hydrophilicity of the former light chain interface by replacing exposed hydrophobic residues with structurally compatible hydrophilic substitutions. The engineered domains can be expressed recombinantly at high yield, are predominantly monomeric at high concentrations, unfold reversibly, and are even more thermostable than typical camelid V(H)H domains. Many of the stabilizing mutations are rare in natural V(H) and V(H)H domains and thus could not be predicted by studying natural sequences and structures. The results demonstrate that autonomous V(H) domains with structural properties beyond the scope of natural frameworks can be derived by using non-natural mutations, which differ from those found in camelid V(H)H domains. These findings should enable the development of libraries of synthetic V(H) domains with CDR3 diversities unconstrained by structural demands. | |||
==About this Structure== | ==About this Structure== | ||
3B9V is a [http://en.wikipedia.org/wiki/Protein_complex Protein complex] structure of sequences from [http://en.wikipedia.org/wiki/Homo_sapiens Homo sapiens]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=3B9V OCA]. | 3B9V is a [http://en.wikipedia.org/wiki/Protein_complex Protein complex] structure of sequences from [http://en.wikipedia.org/wiki/Homo_sapiens Homo sapiens]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=3B9V OCA]. | ||
==Reference== | |||
Comprehensive Analysis of the Factors Contributing to the Stability and Solubility of Autonomous Human VH Domains., Barthelemy PA, Raab H, Appleton BA, Bond CJ, Wu P, Wiesmann C, Sidhu SS, J Biol Chem. 2008 Feb 8;283(6):3639-54. Epub 2007 Nov 28. PMID:[http://ispc.weizmann.ac.il//pmbin/getpm?pmid=18045863 18045863] | |||
[[Category: Homo sapiens]] | [[Category: Homo sapiens]] | ||
[[Category: Protein complex]] | [[Category: Protein complex]] | ||
[[Category: Appleton, B | [[Category: Appleton, B A.]] | ||
[[Category: Barthelemy, P | [[Category: Barthelemy, P A.]] | ||
[[Category: Wiesmann, C.]] | [[Category: Wiesmann, C.]] | ||
[[Category: antibody engineering]] | [[Category: antibody engineering]] | ||
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[[Category: x-ray crystallography]] | [[Category: x-ray crystallography]] | ||
''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 19:04:29 2008'' | ||
Revision as of 20:04, 21 February 2008
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Crystal Structure of an Autonomous VH Domain
OverviewOverview
We report a comprehensive analysis of sequence features that allow for the production of autonomous human heavy chain variable (V(H)) domains that are stable and soluble in the absence of a light chain partner. Using combinatorial phage-displayed libraries and conventional biophysical methods, we analyzed the entire former light chain interface and the third complementarity determining region (CDR3). Unlike the monomeric variable domains of camelid heavy chain antibodies (V(H)H domains), in which autonomous behavior depends on interactions between the hydrophobic former light chain interface and CDR3, we find that the stability of many in vitro evolved V(H) domains is essentially independent of the CDR3 sequence and instead derives from mutations that increase the hydrophilicity of the former light chain interface by replacing exposed hydrophobic residues with structurally compatible hydrophilic substitutions. The engineered domains can be expressed recombinantly at high yield, are predominantly monomeric at high concentrations, unfold reversibly, and are even more thermostable than typical camelid V(H)H domains. Many of the stabilizing mutations are rare in natural V(H) and V(H)H domains and thus could not be predicted by studying natural sequences and structures. The results demonstrate that autonomous V(H) domains with structural properties beyond the scope of natural frameworks can be derived by using non-natural mutations, which differ from those found in camelid V(H)H domains. These findings should enable the development of libraries of synthetic V(H) domains with CDR3 diversities unconstrained by structural demands.
About this StructureAbout this Structure
3B9V is a Protein complex structure of sequences from Homo sapiens. Full crystallographic information is available from OCA.
ReferenceReference
Comprehensive Analysis of the Factors Contributing to the Stability and Solubility of Autonomous Human VH Domains., Barthelemy PA, Raab H, Appleton BA, Bond CJ, Wu P, Wiesmann C, Sidhu SS, J Biol Chem. 2008 Feb 8;283(6):3639-54. Epub 2007 Nov 28. PMID:18045863
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