Data/hora: 14/06/2024 16:00:10

Provedor de dados: | 69
País: | Chile
Título: | Efficient expression and characterization of a cold-active endo-1, 4-&amp;#946;-glucanase from Citrobacter farmeri by co-expression of Myxococcus xanthus protein S
Autores: | Bai,Xi; Yuan,Xianjun; Wen,Aiyou; Li,Junfeng; Bai,Yunfeng; Shao,Tao.
Data: | 2016-11-01
Ano: | 2016
Palavras-chave: | Cellulose degradation; Cellulose; Cold-active enzyme; Endoglucanases; Enzymatic properties; Escherichia coli; Expression; Novel expression vector; N-terminal fusion; Protein S-tag; Recombinant protein.
Resumo: | Background: Cold-active endo-1, 4-&amp;#946;-glucanase (EglC) can decrease energy costs and prevent product denaturation in biotechnological processes. However, the nature EglC from C. farmeri A1 showed very low activity (800 U/L). In an attempt to increase its expression level, C. farmeri EglC was expressed in Escherichia coli as an N-terminal fusion to protein S (ProS) from Myxococcus xanthus. Results: A novel expression vector, pET(ProS-EglC), was successfully constructed for the expression of C. farmeri EglC in E. coli. SDS-PAGE showed that the recombinant protein (ProS-EglC) was approximately 60 kDa. The activity of ProS-EglC was 12,400 U/L, which was considerably higher than that of the nature EglC (800 U/L). ProS-EglC was active at pH 6.5-pH 8.0, with optimum activity at pH 7.0. The recombinant protein was stable at pH 3.5-pH 6.5 for 30 min. The optimal temperature for activity of ProS-EglC was 30°C-40°C. It showed greater than 50% of maximum activity even at 5°C, indicating that the ProS-EglC is a cold-active enzyme. Its activity was increased by Co2+ and Fe2+, but decreased by Cd2+, Zn2+, Li+, methanol, Triton-X-100, acetonitrile, Tween 80, and SDS. Conclusions: The ProS-EglC is promising in application of various biotechnological processes because of its cold-active characterizations. This study also suggests a useful strategy for the expression of foreign proteins in E. coli using a ProS tag.
Tipo: | Journal article
Idioma: | Inglês
Identificador: | http://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0717-34582016000600012
Editor: | Pontificia Universidad Católica de Valparaíso
Formato: | text/html
Fonte: | Electronic Journal of Biotechnology  v.19 n.6 2016