Purification and characterization of a mammalian endo-exonuclease.
An endo-exonuclease has been purified from cultured monkey (CV-1) cells. The enzyme which was purified to near homogeneity to be a 65 kDa monomeric protein. The single-strand DNase activity is endonucleolytic and nonprocessive, whereas the double-strand DNase activity is exonucleolytic and processive. The enzyme was also found to have RNase activity using poly-rA as substrate. The pH optimum for ss-DNase is 8 and for ds-DNase it is 7.5. Both DNase activities require a divalent metal ion (Mg2+, Mn2+, Ca2+, Zn2+) for activity and exhibit the same kinetics of heat inactivation. The purified protein binds to and cleaves a synthetic Holliday junction substrate. The overall enzymatic characteristics of the mammalian protein are very similar to the putative recombination endo-exonucleases purified from Neurospora crassa, Aspergillus nidulans and Saccharomyces cerevisiae.
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Department of Nuclear Medicine, Faculty of Medicine, Université de Sherbrooke, Quebec, Canada.
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D000818:Animals; D002413:Cations, Divalent / Q000378:metabolism; D002461:Cell Line, Transformed; D002499:Centrifugation, Density Gradient; D003429:Cross Reactions; D004247:DNA / Q000378:metabolism*; D004586:Electrophoresis; D004720:Endonucleases / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D005092:Exonucleases / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D000882:Haplorhini; D006863:Hydrogen-Ion Concentration; D011061:Poly A / Q000378:metabolism*
D002413:Cations, Divalent; D011061:Poly A; D004247:DNA; D004720:Endonucleases; D005092:Exonucleases; C020104:endoexonuclease