Extracellular nuclease from a thermophile, Streptomyces thermonitrificans: purification and characterization of the deoxyribonuclease activity.
An extracellular nuclease from Streptomyces thermonitrificans (designated as nuclease Stn alpha) was purified to homogeneity with an overall yield of 2.8%. The Mr of the purified enzyme was 39.6 kDa. The purified enzyme showed an exclusive requirement of Mn2+ for its activity but is not a metalloprotein. The optimum pH for ds- and ssDNA hydrolysis were 7.0 and 7.5 whereas, the optimum temperature was 40 and 45 degrees C, respectively. The enzyme was inhibited by divalent cations, inorganic phosphate and pyrophosphate but not by 3' and 5' mononucleotides. Nuclease Stn alpha is a multifunctional enzyme and its substrate specificity is in the order of dsDNA>ssDNA>>RNA. The end products of both ds- and ssDNA hydrolysis were predominantly oligonucleotides (80-85%) and a small amount of 3' mononucleotides (10-15%) suggesting an endo mode of action.
Division of Biochemical Sciences, National Chemical Laboratory, Pune 411008, India.
Sumedha S
SS
Deshmukh
Vepatu
V
Shankar
D003851:Deoxyribonucleases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004795:Enzyme Stability; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D008670:Metals / Q000494:pharmacology; D013302:Streptomyces / Q000201:enzymology*; D013379:Substrate Specificity; D013696:Temperature
D004791:Enzyme Inhibitors; D008670:Metals; D003851:Deoxyribonucleases