Characterization of the Bacillus subtilis bacteriophage PBS2-induced DNA polymerase and its associated exonuclease activity.
The DNA polymerase induced by Bacillus subtilis bacteriophage PBS2 has a Stokes radius of 7.2 in buffers of high ioninc strength, suggesting a molecular weight in the range 145,000 to 195,000. The polypeptide bands observed on gel electrophoresis in dodecyl sulfate have apparent molecular weights of 78,000 and 69,000 (and possibly another 27,000) in equimolar amounts. In buffers of low ionic strength, the enzyme appears to form large aggregates and even precipitates, with about 90% loss of activity. A nuclease activity co-purifies with the PBS2 DNA polymerase and shows similar responses to changes in pH, MgCl2, N-ethylmaleimide, temperature, and dextran sulfate levels. The nuclease produces deoxyribonucleoside 5'monophosphates from denatured DNA containing thymine or uracil. No endonuclease activity is detectable on supercoiled DNA. The inhibition of nuclease activity by added deoxyribonucleoside triphosphates, the DNA-dependent turnover of triphosphates, to free monophosphates during DNA polymerization, the inhibition of nuclease activity by 3'-phosphates on the DNA template-primer, and the pattern of digestion of 5'-[32P]phosphate-labeled DNA all indicate that the PBS2 DNA polymerase-associated hydrolytic activity is a 3' leads to 5'-exonuclease.
D001412:Bacillus subtilis / Q000201:enzymology*; D001435:Bacteriophages / Q000201:enzymology*; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism*; D003851:Deoxyribonucleases / Q000378:metabolism*; D004790:Enzyme Induction; D005092:Exonucleases / Q000378:metabolism*; D007700:Kinetics; D046911:Macromolecular Substances; D008970:Molecular Weight; D011487:Protein Conformation
D046911:Macromolecular Substances; D004259:DNA-Directed DNA Polymerase; D003851:Deoxyribonucleases; D005092:Exonucleases