An extracellular endodeoxyribonuclease from Streptomyces aureofaciens.
Several extracellular DNases were detected after cultivation of Streptomyces aureofaciens B96 under submerged conditions. These DNases are nutritionally regulated and high content of amino acid nitrogen in cultivation medium repress their production. By varying cultivation conditions, there remained only two extracellular nuclease activities. The major one, extracellular endodeoxyribonuclease SaD I, was purified to homogeneity by ammonium sulfate precipitation, adsorption on Spheron, chromatography on Superose-12P followed by FPLC on MonoQ and final purification on HiTrapQ. The molecular weight of the purified SaD I determined by SDS-PAGE was 31 kDa. The DNase hydrolyses endonucleolytically both double-stranded and single-stranded circular and linear DNA. It does not cleave RNA and does not exhibit phosphodiesterase nor phosphomonoesterase activity. It requires a divalent cation (Zn2+, Co2+, Mn2+, Mg2+) and its activity optimum is at neutral pH (pH 7.2). The optimal temperature for DNA cleavage was 40 degrees C. Activity was strongly inhibited in the presence of phosphate, Hg2+, chelating agents or iodoacetate, but it was stimulated by addition of dimethyl sulphoxide.
Brnáková
Zuzana
Z
Institute of Molecular Biology, Member of the Centre of Excellence for Molecular Medicine, Slovak Academy of Sciences, Dúbravská cesta 21, SK-84551 Bratislava, Slovakia. zuzana.brnakova@savba.sk
Godány
Andrej
A
Timko
Jozef
J
D004706:Endodeoxyribonucleases / Q000302:isolation & purification* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D013304:Streptomyces aureofaciens / Q000201:enzymology*; D013379:Substrate Specificity; D013696:Temperature; D015032:Zinc / Q000494:pharmacology
D004706:Endodeoxyribonucleases; D015032:Zinc