Abstract.
The enzyme β-galactosidase was purified from a cold-adapted organism isolated from Antarctica. The organism was identified as a psychrotrophic Pseudoalteromonas sp. The enzyme was purified with high yields by a rapid purification scheme involving extraction in an aqueous two-phase system followed by hydrophobic interaction chromatography and ultrafiltration. The β-galactosidase was optimally active at pH 9 and at 26 °C when assayed with o-nitrophenyl-β-D-galactopyranoside as substrate for 2 min. The enzyme activity was highly sensitive to temperature above 30 °C and was undetectable at 40 °C. The cations Na+, K+, Mg2+ and Mn2+ activated the enzyme while Ca2+, Hg2+, Cu2+ and Zn2+ inhibited activity. The shelf life of the pure enzyme at 4 °C was significantly enhanced in the presence of 0.1% (w/v) polyethyleneimine. The pure β-galactosidase was also evaluated for lactose hydrolysis. More than 50% lactose hydrolysis was achieved in 8 h in buffer at an enzyme concentration of 1 U/ml, and was increased to 70% in the presence of 0.1% (w/v) polyethyleneimine. The extent of lactose hydrolysis was 40–50% in milk. The enzyme could be immobilized to Sepharose via different chemistries with 60–70% retention of activity. The immobilized enzyme was more stable and its ability to hydrolyze lactose was similar to that of the soluble enzyme.
Similar content being viewed by others
Author information
Authors and Affiliations
Additional information
Electronic Publication
Rights and permissions
About this article
Cite this article
Fernandes, .S., Geueke, .B., Delgado, .O. et al. β-Galactosidase from a cold-adapted bacterium: purification, characterization and application for lactose hydrolysis. Appl Microbiol Biotechnol 58, 313–321 (2002). https://doi.org/10.1007/s00253-001-0905-4
Received:
Revised:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/s00253-001-0905-4