Construction of protease resistant mutein of human CNTF and its expression in Pichia pastoris / 生物工程学报
Chinese Journal of Biotechnology
;
(12): 394-397, 2004.
Artigo
em Chinês
| WPRIM
| ID: wpr-249975
ABSTRACT
AX15 is a mutein of naturally occurring human ciliary neurophic factor (hCNTF), with improved biological activity, stability and solubility. AX15 is susceptible to protease degradation when expressed in Pichia pastoris. Amino acid sequencing revealed the degradation was occurred behind position 12 and 13 amino acid residues, which constitute a dibasic site, RR. Based on the substrate specificity of KEX2, a KEX2 resistant mutein of AX15-AX15 (R13K) was constructed, in which RR was replaced by RK. It was demonstrated that the stability of AX15 (R13K) improved significantly, as no degradation was detected even after 120 hours of induction. AX15 (R13K) was purified to homogeneity by ultrafiltration and gel filtration. TF-1 cell survival bioassay showed AX15 (R13K) had equivalent specific activity to AX15. The protease resistant mutein of AX15 may have greater in vivo stability and thus have superior therapeutic potential.
Texto completo:
DisponíveL
Índice:
WPRIM (Pacífico Ocidental)
Assunto principal:
Peptídeo Hidrolases
/
Pichia
/
Proteínas Recombinantes
/
Química
/
Fator Neurotrófico Ciliar
/
Proteínas Mutantes
/
Vetores Genéticos
/
Genética
/
Metabolismo
/
Mutação
Limite:
Humanos
Idioma:
Chinês
Revista:
Chinese Journal of Biotechnology
Ano de publicação:
2004
Tipo de documento:
Artigo
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