A2 domain of human von Willebrand factor expressed in E. coli and its biological activity / 中国实验血液学杂志
Journal of Experimental Hematology
;
(6): 1090-1093, 2005.
Artigo
em Chinês
| WPRIM
| ID: wpr-343820
ABSTRACT
Von Willebrand factor (vWF) is the unique substrate for the metalloprotease, ADAMTS-13, and plays a pivotal role in the pathology of von Willebrand disease (vWD) and thrombotic thrombocytopenic purpure (TTP). To study the pathogenesis of TTP and to establish a method to diagnose TTP, the DNA fragment of vWF-A2 domain was amplified and inserted into expression vector with 6 x His tag (pQE-30), the recombinant expression vector was transformed into E. coli (strain M15) and induced by IPTG. The recombinant fragment comprising residues 718-905 of mature vWF was designated as rvWF-A2. It was purified by Ni-NTA resin column chromatography and refolded in Tris buffer containing GSH and GSSG. The results demonstrated that rvWF-A2 was expressed successfully in E. coli M15, amounting to 42% of total bacterial protein with the purity over 98%. It was identified that rvWF-A2 can be efficiently cleaved by the citrated normal plasma while no cleavage can be detected by the TTP plasma or plasma with EDTA. It is concluded that rvWF-A2 expressed efficiently in E. coli demonstrated excellent biological activity, which lays a solid foundation for establishment of method to measure quantatively the activity of ADAMTS-13.
Texto completo:
DisponíveL
Índice:
WPRIM (Pacífico Ocidental)
Assunto principal:
Fragmentos de Peptídeos
/
Púrpura Trombocitopênica Trombótica
/
Proteínas Recombinantes
/
Fator de von Willebrand
/
Química
/
Diagnóstico
/
Escherichia coli
/
Proteínas ADAM
/
Proteína ADAMTS13
/
Genética
Tipo de estudo:
Estudo diagnóstico
/
Estudo prognóstico
Limite:
Humanos
Idioma:
Chinês
Revista:
Journal of Experimental Hematology
Ano de publicação:
2005
Tipo de documento:
Artigo
Similares
MEDLINE
...
LILACS
LIS