Expression, purification and characterization of a thermostable lactate dehydrogenase from Thermotoga maritima / 生物工程学报
Chinese Journal of Biotechnology
;
(12): 545-553, 2014.
Artigo
em Chinês
| WPRIM
| ID: wpr-279484
ABSTRACT
The gene encoding thermostable lactate dehydrogenase (Tm-LDH) was cloned into the plasmid pHsh from Thermotoga maritima, and expressed in Escherichia coli JM 109. The recombinant protein was purified to homogeneity by a simple step, heat treatment. The recombinant enzyme had a molecular mass of 33 kDa. The optimal temperature and pH of Tm-LDH were observed 95 degrees C and 7.0. The purified enzyme had a half-life of 2 h at 90 degrees C, and exhibited better stability over a pH range from 5.5 to 8.0. The K(m) and V(max) values were 1.7 mmol/L, 3.8 x 10(4) U/mg of protein for pyruvate, and 7.2 mmol/L and 1.1 x 10(5) U/mg for NADH, respectively. The expression of Tm-LDH in T7 system could not obtain high efficiency, but it has been soluble over-expression in pHsh system and reached 340 mg/L. The superior stability and productivity of Tm-LDH will lay the foundation of its industrial-scale fermentation and application in the NAD regeneration.
Texto completo:
DisponíveL
Índice:
WPRIM (Pacífico Ocidental)
Assunto principal:
Temperatura
/
Estabilidade Enzimática
/
Proteínas Recombinantes
/
Clonagem Molecular
/
Thermotoga maritima
/
Escherichia coli
/
L-Lactato Desidrogenase
/
Metabolismo
/
Peso Molecular
Idioma:
Chinês
Revista:
Chinese Journal of Biotechnology
Ano de publicação:
2014
Tipo de documento:
Artigo
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