Exonuclease III

 

    Exonuclease III is a nuclease that stepwise removes mononucleotides from the 3'-OH termini of double-stranded DNA (dsDNA), and inactive on single-stranded DNA (ssDNA). The preferred substrates of Exonuclease III are blunt or recessed 3'-termini, although the enzyme also acts at nick sites in double-stranded DNA to generate single-stranded gaps.3'-overhanging termini are resistant to cleavage; the degree of resistance depends on the length of the extension, with extensions 4 bases or longer being essentially resistant to cleavage. This characteristic can be used to produce strand-specific ssDNA with designed linearized dsDNA having one non-cleavage end (3'-overhang) and the other end designed as a cleavable end (blunt or 5'-overhang), allowing Exonuclease III to digest only one strand.
    Exonuclease III also exhibits RNase H, 3'-phosphatase, and apurinic/apyrimidinic-endonuclease activities.