Identification and purification of a single-stranded-DNA-specific exonuclease encoded by the recJ gene of Escherichia coli.
Abstract
The Escherichia coli recJ gene product was overproduced using a plasmid that carries the recJ gene downstream of a strong regulatable promoter and a strong ribosome-binding site. Overexpression of recJ produced a concomitant increase in the levels of single-stranded-DNA-specific nuclease activity present in crude cell extracts. This nuclease activity was purified to homogeneity and found to reside in a 60-kDa polypeptide. This polypeptide was induced with recJ overexpression and had the size and N-terminal amino acid sequence identical to the predicted RecJ protein sequence. The RecJ nuclease degraded linear single-stranded DNA but did not have exonuclease activity on linear double-stranded substrates or endonuclease activity on either single-stranded or double-stranded substrates. The RecJ exonuclease had greater activity on duplex DNA molecules with 5'-rather than 3'-single-stranded tails.
- Publication:
-
Proceedings of the National Academy of Science
- Pub Date:
- April 1989
- DOI:
- 10.1073/pnas.86.8.2627
- Bibcode:
- 1989PNAS...86.2627L