| Summary: | National Infrastructure of Natural Resources for Science and Technology Program of China [2005DKA21209]; National High-Tech Program [2007AA091904]; China Ocean Mineral Resources RD Association [COMRA DYXM-115-02-2-03] A psychrotrophic bacterium, Arthrobacter sp. ON14, isolated from Antarctica, was shown to exhibit a high beta-galactosidase activity at a low temperature. A genomic library of ON14 was constructed and screened for beta-galactosidase genes on functional plates containing 5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside (X-gal) as the substrate. Two different beta-galactosidase genes, named as galA, gal B, were found in ON14. Computational analyses of the genes revealed that the encoded protein GalA belongs to family 2 of glycosyl hydrolysases and is a cold-active protein, whereas GalB belongs to family 42 of glycosyl hydrolysases and is a mesophilic protein. Reverse transcription analyses revealed that the expression of galA is highly induced at a low temperature (4 degrees C) and repressed at a high temperature (28 degrees C) when lactose is used as the sole carbon source. Conversely, the expression of galB is inhibited at a low temperature and induced at a high temperature. The purified GalA showed its peak activity at 15 degrees C and pH 8. The mineral ions Na(+), K(+), Mg(2+), and Mn(2+) were identified as enzyme activators, whereas Ca(2+) had no influence on the enzyme activity. An enzyme stability assay revealed that the activity of GalA is significantly decreased when it is incubated at 45 degrees C for 2 h, and all its activity is lost when it is incubated at 50 degrees C.
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