Dissociation of alpha beta DNA polymerase of avian myeloblastosis virus by dimethyl sulfoxide.
The alpha beta DNA polymerase of avian myeloblastosis virus was treated with dimethyl sulfoxide to dissociate the enzyme subunits. The dimethyl sulfoxide treated enzymes were passed over phosphocellulose to purify and characterize the dissociated subunits as well as to remove the dimethyl sulfoxide. RNA-directed DNA polymerase, RNase H, and nucleic acid-binding activity were monitored, as well as the subunit structure (on sodium dodecyl sulfate-polyacrylamide gels) of the various enzyme species obtained. With 30% dimethyl sulfoxide, the majority of DNA polymerase and RNase H activities as well as the alpha subunit were displaced from the alpha beta DNA polymerase position on phosphocellulose (0.23 M potassium phosphate) to the alpha DNA polymerase position (0.1 M). The association of DNA polymerase and RNase H activities with the alpha subunit suggests that alpha is the enzymatically active subunit in alpha beta. In addition to alpha DNA polymerase, a minor polymerase species eluted from phosphocellulose at 0.4 M potassium phosphate. The dissociated beta subunit eluted from phosphocellulose at a wide range of salt concentrations (0.28 to 0.5 M potassium phosphate). The dissociated beta subunit bound 3H-labeled murine leukemia virus RNA and [3H]poly(dT)-poly(dA) approximately 20-fold more avidly than alpha DNA polymerase alone. In contrast to the results with the alpha subunit, there was no correlation between DNA polymerase and RNase H activity profiles and the elution profile of the beta subunit from phosphocellulose. These observations suggest the beta subunit is either enzymatically inactive or possesses limited DNA polymerase and RNase H activity when compared with the alpha subunit.
D001354:Avian Leukosis Virus / Q000201:enzymology*; D009189:Avian Myeloblastosis Virus / Q000201:enzymology*; D004279:DNA, Viral / Q000096:biosynthesis; D004121:Dimethyl Sulfoxide / Q000494:pharmacology*; D011061:Poly A / Q000378:metabolism; D011071:Poly T / Q000378:metabolism; D011485:Protein Binding; D012194:RNA-Directed DNA Polymerase / Q000302:isolation & purification / Q000378:metabolism*; D012260:Ribonucleases / Q000378:metabolism; D013698:Templates, Genetic
D004279:DNA, Viral; D011061:Poly A; D011071:Poly T; D012194:RNA-Directed DNA Polymerase; D012260:Ribonucleases; D004121:Dimethyl Sulfoxide