Purification and characterization of a cold-adapted uracil-DNA glycosylasefrom Atlantic cod (Gadus morhua)

Uracil-DNA glycosylase (UDG; UNG) has been purified 17000-fold from Atlanti c cod liver (Gadus,,morhua). The enzyme has an apparent molecular mass of 2 5 kDa, as determined by gel filtration, and an isoelectric point above 9.0. Atlantic cUNG is inhibited by the specific UNG inhibitor (Ugi) from the Ba cillus subtilis bacteriophage (PBS?), and has a 2-fold higher activity for single-stranded DNA than for double-stranded DNA. cUNG has an optimum activ ity between pll 7.0-9.0 and 25 50 mM NaCl, and a temperature optimum of 41 degreesC. Cod UNG was compared with the recombinant human UNG (rhUNG), and was found to have slightly higher relative activity at low temperatures com pared with their respective optimum temperatures. Cod UNG is also more pH- and temperature labile than rhUNG. At pll 10.0, the recombinant human UNG h ad 66% residual activity compared with only 0.4% for the Atlantic cUNG. At 50 degreesC, cUNG had a half-life of 0.5 min compared with 8 min for the rh UNG. These activity and stability experiments reveal cold-adapted features in cUNG. (C) 2000 Elsevier Science Inc. All rights reserved.