ABSTRACT
In the current investigation, aqueous two-phase methodology with polyethylene glycol (PEG) 6000/phosphate salt has been used for single-step purification of β-galactosidase from recombinant Pichia pastoris. Optimized parameters with 12% (w/w) salt concentration, 25% (w/w) polymer concentration, 0.6 (mg/ml) protein load and 0.1 M ionic concentration resulted in a maximum of 4.7 purification fold with a 97% yield. The enzyme kinetic study of purified protein revealed Vm and Km of 97.087 (U/mg), 0.027(U/mg), 0.07 (U/mg) and 0.7 (mM), 11.13 (mM), 10.73 (mM) with O-nitrophenyl-β-D-galactopyranoside, lactose and milk substrate, respectively.
Acknowledgments
We are grateful to the Department of Biosciences and Bioengineering, Indian Institute of Technology Guwahati, Guwahati, Assam, India for providing facilities to carry out this research.
Conflict of interest
The authors have declared no conflict of interest.