Purification of E. coli-synthesized Pan proteins and development of a Pan-specific monoclonal antibody.
Hybridoma
; 13(3): 191-7, 1994 Jun.
Article
em En
| MEDLINE
| ID: mdl-7523277
ABSTRACT
The helix-loop-helix (HLH) transcription factors, Pan-1 (E47) and Pan-2 (E12), are produced by the mechanism of alternative transcript splicing. Pan-1 and Pan-2 were expressed in Escherichia coli, and a purification scheme was developed. Purified Pan-2 was used to immunize Smith-Webster mice and a hybridoma was generated that produced a monoclonal antibody (Yae) that specifically recognized both native and denatured Pan-1 and Pan-2. Deletion mapping and sequence transfer studies have localized the determinant recognized by the Yae antibody to the region 195-208 of Pan-2. This region is conserved in Pan-1 and Pan-2. The Yae antibody recognized in vitro-synthesized ITF-1, a third E2A (Pan) gene product also produced by the mechanism of alternative RNA splicing, but did not recognize the related HLH proteins, ITF-2, REB alpha, or REB beta. By Western blot assay of pancreatic acinar cells, the Yae antibody detected a single protein species of 72 kD that comigrated with in vitro-synthesized Pan-1 and Pan-2.
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Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Fatores de Transcrição
/
Proteínas de Ligação a DNA
/
Anticorpos Monoclonais
Limite:
Animals
Idioma:
En
Ano de publicação:
1994
Tipo de documento:
Article