Gene cloning and soluble expression of Aspergillus niger phytase in E. coli cytosol via chaperone co-expression.
A phytase gene from Aspergillus niger was isolated and two Escherichia coli expression systems, based on T7 RNA polymerase promoter and tac promoter, were used for its recombinant expression. Co-expression of molecular chaperone, GroES/EL, aided functional cytosolic expression of the phytase in E. coli BL21 (DE3). Untagged and maltose-binding protein-tagged recombinant phytase showed an activity band of ~49 and 92 kDa, respectively, on a zymogram. Heterologously-expressed phytase was fractionated from endogenous E. coli phytase by (NH4)2SO4 precipitation. The enzyme had optimum activity at 50 °C and pH 6.5.
Mrudula Vasudevan
MV
Ushasree
Jalaja
J
Vidya
Ashok
A
Pandey
D010832:6-Phytase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001234:Aspergillus niger / Q000201:enzymology* / Q000235:genetics; D003001:Cloning, Molecular; D003600:Cytosol / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006031:Glycosylation; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D058619:Maltose-Binding Proteins; D018832:Molecular Chaperones / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013696:Temperature
D005656:Fungal Proteins; D058619:Maltose-Binding Proteins; D018832:Molecular Chaperones; D011994:Recombinant Proteins; D010832:6-Phytase