Secretory expression and characterization of heat sensitive nuclease in Pichia pastoris / 生物工程学报
Chinese Journal of Biotechnology
;
(12): 991-995, 2016.
Artigo
em Chinês
| WPRIM
| ID: wpr-242280
ABSTRACT
Nucleases is an important enzyme widely used in biotechnology. A codon optimized nuclease gene (SNU) from Northern Shrimps was inserted into pPICZα A vector, and expressed extracellularly in strain SMD1168H. On the basis of multi-copy recombinant strain, we further optimized the expression condition and characterized SNU. SNU was highly expressed and stable after 1% methanol induction for 72 h, yield reached 1.4×10⁵ U/mL. SDS-PAGE electrophoresis demonstrated that this is a N-linked glycoprotein of 50 kDa. It was purified by one step DEAE Sephadex chromatography to the purity of about 15 mg/L with a specific activity of 6.291×10⁶ U/mg. Functional analysis on the nuclease activity indicated that it was stimulated by bivalent iron, such as Ca²⁺, Mn²⁺, Co²⁺ and Mg²⁺, but inhibited by Zn²⁺, Cu²⁺ and high salt. Meanwhile, it was irreversibly inactivated at 70 ℃ for 10 min.
Texto completo:
DisponíveL
Índice:
WPRIM (Pacífico Ocidental)
Assunto principal:
Pichia
/
Proteínas Recombinantes
/
Códon
/
Glicoproteínas
/
Penaeidae
/
Eletroforese em Gel de Poliacrilamida
/
Endonucleases
/
Temperatura Alta
/
Metabolismo
Tipo de estudo:
Estudo diagnóstico
Limite:
Animais
Idioma:
Chinês
Revista:
Chinese Journal of Biotechnology
Ano de publicação:
2016
Tipo de documento:
Artigo
Similares
MEDLINE
...
LILACS
LIS