Construction and expression of protein self-splicing prokaryotic expression vector pTWIN1- AcAPc2 / 生物医学工程学杂志
Journal of Biomedical Engineering
;
(6): 630-634, 2006.
Artículo
en Chino
| WPRIM
| ID: wpr-249541
ABSTRACT
To express recombinant Ancylostoma caninum anticoagulant peptide-c2 (AcAPc2), a whole cDNA fragment encoding AcAPc2 was achieved by ligation- PCR and inserted into prokaryotic expression vector pTWIN1 for constructing the specific self-splicing prokaryotic expression vector, pTWIN1-AcAPc2; positive recombinants were transformed into E. coli ER2566 for expression research. The recombinant protein, AcAPc2-intein2-CBD, was soluble and expressed in E. coli ER2566 (about 30.1% fusion protein in total protein). AcAPc2-intein2-CBD was characterized to be 41 KD by SDS-PAGE and identified by Western-blot. The recombinant fusion protein was purified to a efficiently high degree by chitin affinity chromatography. After the process of specific self-splicing induced by beta-Mercaptoethanol, the target protein, AcAPc2, was obtained, characterized to be 21 KD by SDS-PAGE and migrated as a dimmer. Molecular weight of AcAPc2 conformed to native dimmer. Bio-information analysis indicated relationship between secondary construction of AcAPc2 and biologic function. These findings greatly facilitate the purification of AcAPc2 and are very important for the additional studies on its anti-coagulation mechanism and its clinical application as anti-coagulation medicine.
Texto completo:
Disponible
Índice:
WPRIM (Pacífico Occidental)
Asunto principal:
Farmacología
/
Plásmidos
/
Células Procariotas
/
Proteínas Recombinantes de Fusión
/
Expresión Génica
/
Proteínas del Helminto
/
Química
/
Empalme del ARN
/
Genes de Helminto
/
Escherichia coli
Tipo de estudio:
Estudio pronóstico
Límite:
Animales
Idioma:
Chino
Revista:
Journal of Biomedical Engineering
Año:
2006
Tipo del documento:
Artículo
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