Construction, expression and characterization of recombinant fusion protein HSA-PTH (1-34) in Pichia pastoris / 浙江大学学报·医学版
Journal of Zhejiang University. Medical sciences
; (6): 126-133, 2008.
Artigo
em Chinês
| WPRIM (Pacífico Ocidental)
| ID: wpr-344364
Biblioteca responsável:
WPRO
ABSTRACT
<p><b>OBJECTIVE</b>To obtain recombinant fusion protein HSA (human serum albumin)-PTH(1-34) in Pichia pastoris.</p><p><b>METHODS</b>HSA and PTH(1-34) cDNA were obtained with PCR and the DNA segments were cloned into vector pPIC9 with linker. The linearized plasmids were transformed GS115 competent cells treated with LiCl, and mut+ transformants were screened on MD plate. With AOX promoter and alpha-MF signal sequences leading, fusion protein was expressed in GS115. PCR and SDS-PAGE were employed to confirm the integration and expression of HSA-PTH(1-34). The fusion protein was identified by Western blotting and classical adenylate cyclase assay.</p><p><b>RESULT</b>The PCR results showed that the gene of HSA-PTH(1-34) was integrated into GS115 genome. Western bolt approved the existence of two domains of HSA and PTH(1-34). The bioactivity assay in rabbit cortical membranes indicated that HSA-PTH (1-34) activated adenylate cyclase, but the activity was lower than that of the synthetic PTH(1-34).</p><p><b>CONCLUSION</b>Active fusion protein HSA-PTH (1-34) is successfully expressed in Pichia pastoris.</p>
Texto completo:
Disponível
Contexto em Saúde:
Doenças Negligenciadas
Problema de saúde:
Doenças Negligenciadas
/
Zoonoses
Base de dados:
WPRIM (Pacífico Ocidental)
Assunto principal:
Fragmentos de Peptídeos
/
Pichia
/
Proteínas Recombinantes de Fusão
/
Albumina Sérica
/
Teriparatida
/
Escherichia coli
/
Vetores Genéticos
/
Genética
/
Metabolismo
Limite:
Humanos
Idioma:
Chinês
Revista:
Journal of Zhejiang University. Medical sciences
Ano de publicação:
2008
Tipo de documento:
Artigo