The Enzyme and the cDNA Sequence of a Thermolabile and Double-Strand Specific DNase from Northern

• Main Authors: Shrimps (pandalus Borealis, Inge W. Nilsen, Kersti Øverbø, Linda Jensen Havdalen, Morten Elde, Dag Rune Gjellesvik
• Other Authors: The Pennsylvania State University CiteSeerX Archives
• Format: Text
• Language: English
• Subjects: Kamchatka; Kamchatka crab; northern shrimp; Pandalus borealis
• Online Access: http://citeseerx.ist.psu.edu/viewdoc/summary?doi=10.1.1.354.1996

Background: We have previously isolated a thermolabile nuclease specific for double-stranded DNA from industrial processing water of Northern shrimps (Pandalus borealis) and developed an application of the enzyme in removal of contaminating DNA in PCR-related technologies. Methodology/Principal Findings: A 43 kDa nuclease with a high specific activity of hydrolysing linear as well as circular forms of DNA was purified from hepatopancreas of Northern shrimp (Pandalus borealis). The enzyme displayed a substrate preference that was shifted from exclusively double-stranded DNA in the presence of magnesium to also encompass significant activity against single-stranded DNA when calcium was added. No activity against RNA was detected. Although originating from a cold-environment animal, the shrimp DNase has only minor low-temperature activity. Still, the enzyme was irreversibly inactivated by moderate heating with a half-life of 1 min at 65uC. The purified protein was partly sequenced and derived oligonucleotides were used to prime amplification of the encoding cDNA. This cDNA sequence revealed an open reading frame encoding a 404 amino acid protein containing a signal peptide. By sequence similarity the enzyme is predicted to belong to a family of DNA/RNA non-specific nucleases even though this shrimp DNase lacks RNase activity and is highly double-strand specific in some respects. These features are in agreement with those previously established for endonucleases classified as similar to the Kamchatka crab duplex-specific nuclease (Par_DSN). Sequence comparisons and