Isolation and some properties of two deoxyribonucleases from a snail, Achatina fulica.
1. Two main DNases were found in the dried liver extract of a snail, Achatina fulica. They were purified by the phosphocellulose batch method and by phosphocellulose column chromatography. The enzyme eluted earlier from the phosphocellulose column was designated as Achatina DNase-1 and the other as Achatina DNase-2. DNase-1 was purified further by QAE-Sephadex A-25 column chromatography (twice) just before use because of the instability of the purified enzyme. By these procedures, DNase-1 and 2 were purified 200- and 130-fold, respectively. 2. Divalent or monovalent cations had no marked effect on either enzyme. The showed pH optima of 4.8 (DNase-1) and 5.2 (DNase-2). Ionic strength was found to be critical for the maximal activity. The isoelectric points of DNase-1 and 2 were both 6.9. On heating at 70--75 degrees C for 5 min, each enzymic activity fell to half of the initial value. 3. The enzyme preparations degraded native DNA 1.5--2.5 times faster than heat-denatured DNA. They both degraded heat-denatured DNA endonucleolytically, to give oligonucleotides with 3'-phosphates. 4. The 3'-phosphoryl and 5'-hydroxy termini of the resulting oligonucleotides were analyzed. DNase-1 possessed marked specificity for dThd at 3'-termini and dAdo at 5'-termini in the early stages of degradation, but only for dAdo at 5'-termini in the later stages. DNase-2 showed some preference for purine nucleotides at both 3'- and 5'-termini in the later stages of degradation.
D002845:Chromatography / Q000379:methods; D002852:Chromatography, Ion Exchange / Q000379:methods; D004247:DNA / Q000378:metabolism; D004277:DNA, Single-Stranded / Q000378:metabolism; D003851:Deoxyribonucleases / Q000302:isolation & purification* / Q000378:metabolism; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D008099:Liver / Q000201:enzymology*; D009691:Nucleic Acid Denaturation; D012908:Snails / Q000201:enzymology*; D013379:Substrate Specificity
D004277:DNA, Single-Stranded; D004247:DNA; D003851:Deoxyribonucleases