Aggregation-resistant domain antibodies selected on phage by heat denaturation.
Nat Biotechnol
; 22(9): 1161-5, 2004 Sep.
Article
em En
| MEDLINE
| ID: mdl-15300256
ABSTRACT
We describe a method for selecting aggregation-resistant proteins by heat denaturation. This is illustrated with antibody heavy chain variable domains (dAbs), which are prone to aggregate. The dAbs were displayed multivalently at the infective tip of filamentous bacteriophage, and heated transiently to induce unfolding and to promote aggregation of the dAbs. After cooling, the dAbs were selected for binding to protein A (a ligand common to these folded dAbs). Phage displaying dAbs that unfold reversibly were thereby enriched with respect to those that do not. From a repertoire of phage dAbs, six dAbs were characterized after selection; they all resisted aggregation, and were soluble, well expressed in bacteria and could be purified in good yields. The method should be useful for making aggregation-resistant proteins and for helping to identify features that promote or prevent protein aggregation, including those responsible for misfolding diseases.
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Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Região Variável de Imunoglobulina
/
Ensaio de Imunoadsorção Enzimática
/
Cadeias Pesadas de Imunoglobulinas
/
Complexos Multiproteicos
/
Temperatura Alta
Tipo de estudo:
Evaluation_studies
/
Prognostic_studies
Idioma:
En
Revista:
Nat Biotechnol
Assunto da revista:
BIOTECNOLOGIA
Ano de publicação:
2004
Tipo de documento:
Article