Isolation and identification of a strain producing cold-adapted beta-galactosidase, and purification and characterisation of the enzyme

Enzymes with high specific activities at low temperatures have potential uses in the food industry. Cold-adapted microorganisms are potentially useful sources of cold-active enzyme. To find cold-adapted beta-galactosidase, we isolated several cold-adapted microorganisms from glacier zone soil. One cold-adapted beta-galactosidase producing strain was obtained. The biochemical characteristics and the results of 16S rDNA sequencing identified the strain as Rahnella aquatilis. The enzyme was purified by column chromatography, after which a single protein band migrating near 60 kDa was observed by means of SDS-PAGE. The beta-galactosidase was optimally active at 35 deg C and at pH 6.5 when assayed with o-nitrophenyl-beta-D-galactopyranoside as substrate. The enzyme activity was sensitive to temperatures above 40 deg C and was undetectable at 45 deg C. Metal ions Mn2+ and K+ activated the enzyme, while Cu2+, Zn2+, Fe3+, and Al3+ inhibited the activity. The enzyme was also assayed for lactose hydrolysis. When milk is treated with the enzyme at 30 deg C for 2 h, the degree of lactose hydrolysis can reach 80%. Thus, it has potential applications in the food industry.