Efficient expression and characterization of a cold-active endo-1, 4-β-glucanase from Citrobacter farmeri by co-expression of Myxococcus xanthus protein S
Background: Cold-active endo-1, 4-β-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...
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Pontificia Universidad Católica de Valparaíso
2016
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oai:scielo:S0717-345820160006000122017-01-25Efficient expression and characterization of a cold-active endo-1, 4-β-glucanase from Citrobacter farmeri by co-expression of Myxococcus xanthus protein SBai,XiYuan,XianjunWen,AiyouLi,JunfengBai,YunfengShao,Tao Cellulose degradation Cellulose Cold-active enzyme Endoglucanases Enzymatic properties Escherichia coli Expression Novel expression vector N-terminal fusion Protein S-tag Recombinant protein Background: Cold-active endo-1, 4-β-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.info:eu-repo/semantics/openAccessPontificia Universidad Católica de ValparaísoElectronic Journal of Biotechnology v.19 n.6 20162016-11-01text/htmlhttp://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0717-34582016000600012en10.1016/j.ejbt.2016.10.005 |
| institution |
Scielo Chile |
| collection |
Scielo Chile |
| language |
English |
| topic |
Cellulose degradation Cellulose Cold-active enzyme Endoglucanases Enzymatic properties Escherichia coli Expression Novel expression vector N-terminal fusion Protein S-tag Recombinant protein |
| spellingShingle |
Cellulose degradation Cellulose Cold-active enzyme Endoglucanases Enzymatic properties Escherichia coli Expression Novel expression vector N-terminal fusion Protein S-tag Recombinant protein Bai,Xi Yuan,Xianjun Wen,Aiyou Li,Junfeng Bai,Yunfeng Shao,Tao Efficient expression and characterization of a cold-active endo-1, 4-β-glucanase from Citrobacter farmeri by co-expression of Myxococcus xanthus protein S |
| description |
Background: Cold-active endo-1, 4-β-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. |
| author |
Bai,Xi Yuan,Xianjun Wen,Aiyou Li,Junfeng Bai,Yunfeng Shao,Tao |
| author_facet |
Bai,Xi Yuan,Xianjun Wen,Aiyou Li,Junfeng Bai,Yunfeng Shao,Tao |
| author_sort |
Bai,Xi |
| title |
Efficient expression and characterization of a cold-active endo-1, 4-β-glucanase from Citrobacter farmeri by co-expression of Myxococcus xanthus protein S |
| title_short |
Efficient expression and characterization of a cold-active endo-1, 4-β-glucanase from Citrobacter farmeri by co-expression of Myxococcus xanthus protein S |
| title_full |
Efficient expression and characterization of a cold-active endo-1, 4-β-glucanase from Citrobacter farmeri by co-expression of Myxococcus xanthus protein S |
| title_fullStr |
Efficient expression and characterization of a cold-active endo-1, 4-β-glucanase from Citrobacter farmeri by co-expression of Myxococcus xanthus protein S |
| title_full_unstemmed |
Efficient expression and characterization of a cold-active endo-1, 4-β-glucanase from Citrobacter farmeri by co-expression of Myxococcus xanthus protein S |
| title_sort |
efficient expression and characterization of a cold-active endo-1, 4-β-glucanase from citrobacter farmeri by co-expression of myxococcus xanthus protein s |
| publisher |
Pontificia Universidad Católica de Valparaíso |
| publishDate |
2016 |
| url |
http://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0717-34582016000600012 |
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