Purification and properties of an endonuclease from nuclei of uninfected and polyoma-infected 3T3 cells.
An endonuclease activity has been purified approximately 800-fold from nuclei of 3T3 cells infected with polyoma virus. The purfied enzyme catalyzes an endonucleoytic cleavage of single- and double-stranded DNA and single-stranded RNA. Evidence that the activity towards these substrates resides in the same protein molecule is provided by the finding that they co-sediment in sucrose gradients and have identical rates of heat inactivation. Studies on the DNase activity shows that the rate of hydrolysis of single-stranded T7 DNA is 100-fold greater than that for double-stranded T7 DNA. Single-stranded DNA is extensively hydrolyzed to low molecular weight acid-insoluble products. With duplex DNA as substrate, only a limited number of single strand breaks are introduced. A limit digest with polyoma DNA (component I) as substrate results in the introduction of four breaks per strand. The phosphdiester bond interruptions can be repaired by polynucleotide ligase. Approximately 80% of the 5' termini present at the point of phosphodiester bond cleavage are purine nucleotides. Additional studies have demonstrated that a similar endonuclease is present in nuclei of uninfected cells and that this enzyme purified 400-fold has catalytic properties identical with those of the endonuclease from infected cells.
D002460:Cell Line; D002467:Cell Nucleus / Q000201:enzymology*; D002471:Cell Transformation, Neoplastic; D004277:DNA, Single-Stranded; D003851:Deoxyribonucleases / Q000302:isolation & purification / Q000378:metabolism; D004720:Endonucleases / Q000302:isolation & purification / Q000378:metabolism; D007700:Kinetics; D008970:Molecular Weight; D011120:Polyomavirus / Q000201:enzymology*; D012260:Ribonucleases / Q000302:isolation & purification / Q000378:metabolism
D004277:DNA, Single-Stranded; D003851:Deoxyribonucleases; D004720:Endonucleases; D012260:Ribonucleases