Source:http://linkedlifedata.com/resource/pubmed/id/20543007
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| Predicate | Object |
|---|---|
| rdf:type | |
| lifeskim:mentions | |
| pubmed:issue |
8
|
| pubmed:dateCreated |
2010-7-8
|
| pubmed:abstractText |
Protein aggregation is of great concern to pharmaceutical formulations and has been implicated in several diseases. We engineered an anti-IL-13 monoclonal antibody CNTO607 for improved solubility. Three structure-based engineering approaches were employed in this study: (i) modifying the isoelectric point (pI), (ii) decreasing the overall surface hydrophobicity and (iii) re-introducing an N-linked carbohydrate moiety within a complementarity-determining region (CDR) sequence. A mutant was identified with a modified pI that had a 2-fold improvement in solubility while retaining the binding affinity to IL-13. Several mutants with decreased overall surface hydrophobicity also showed moderately improved solubility while maintaining a similar antigen affinity. Structural studies combined with mutagenesis data identified an aggregation 'hot spot' in heavy-chain CDR3 (H-CDR3) that contains three residues ((99)FHW(100a)). The same residues, however, were found to be essential for high affinity binding to IL-13. On the basis of the spatial proximity and germline sequence, we reintroduced the consensus N-glycosylation site in H-CDR2 which was found in the original antibody, anticipating that the carbohydrate moiety would shield the aggregation 'hot spot' in H-CDR3 while not interfering with antigen binding. Peptide mapping and mass spectrometric analysis revealed that the N-glycosylation site was generally occupied. This variant showed greatly improved solubility and bound to IL-13 with affinity similar to CNTO607 without the N-linked carbohydrate. All three engineering approaches led to improved solubility and adding an N-linked carbohydrate to the CDR was the most effective route for enhancing the solubility of CNTO607.
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| pubmed:language |
eng
|
| pubmed:journal | |
| pubmed:citationSubset |
IM
|
| pubmed:chemical | |
| pubmed:status |
MEDLINE
|
| pubmed:month |
Aug
|
| pubmed:issn |
1741-0134
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| pubmed:author |
pubmed-author:BakerAudreyA,
pubmed-author:CanzianiGabrielaG,
pubmed-author:FengYiqingY,
pubmed-author:GillilandGary LGL,
pubmed-author:HuangMaggieM,
pubmed-author:JacobsSteven ASA,
pubmed-author:KangJamesJ,
pubmed-author:LacyEilyn RER,
pubmed-author:LuoJinquanJ,
pubmed-author:O'NeilKaryn TKT,
pubmed-author:RajuT ShanthaTS,
pubmed-author:TangQing MikeQM,
pubmed-author:TeplyakovAlexeyA,
pubmed-author:WuSheng-JiunSJ
|
| pubmed:issnType |
Electronic
|
| pubmed:volume |
23
|
| pubmed:owner |
NLM
|
| pubmed:authorsComplete |
Y
|
| pubmed:pagination |
643-51
|
| pubmed:dateRevised |
2010-11-18
|
| pubmed:meshHeading |
pubmed-meshheading:20543007-Amino Acid Sequence,
pubmed-meshheading:20543007-Antibodies, Monoclonal,
pubmed-meshheading:20543007-Binding Sites,
pubmed-meshheading:20543007-Calorimetry, Differential Scanning,
pubmed-meshheading:20543007-Electrophoresis, Polyacrylamide Gel,
pubmed-meshheading:20543007-Humans,
pubmed-meshheading:20543007-Hydrophobic and Hydrophilic Interactions,
pubmed-meshheading:20543007-Interleukin-13,
pubmed-meshheading:20543007-Isoelectric Focusing,
pubmed-meshheading:20543007-Isoelectric Point,
pubmed-meshheading:20543007-Models, Molecular,
pubmed-meshheading:20543007-Molecular Sequence Data,
pubmed-meshheading:20543007-Peptide Mapping,
pubmed-meshheading:20543007-Protein Conformation,
pubmed-meshheading:20543007-Protein Engineering,
pubmed-meshheading:20543007-Protein Multimerization,
pubmed-meshheading:20543007-Protein Stability,
pubmed-meshheading:20543007-Solubility,
pubmed-meshheading:20543007-Temperature
|
| pubmed:year |
2010
|
| pubmed:articleTitle |
Structure-based engineering of a monoclonal antibody for improved solubility.
|
| pubmed:affiliation |
Biologics Research, Centocor R&D, 145 King of Prussia Radnor, PA 19087-4557, USA. swu4@its.jnj.com
|
| pubmed:publicationType |
Journal Article,
Research Support, Non-U.S. Gov't
|