DROSOPHILA DNA-POLYMERASE-DELTA - PURIFICATION AND CHARACTERIZATION

A DNA polymerase with properties similar to mammalian polymerase delta has been isolated to near homogeneity from early embryos of Drosophil a melanogaster. A combination of exclusion chromatography and sodium d odecyl sulfate-polyacrylamide gel electrophoresis indicates that this enzyme has a total molecular mass of 185 kDa and is composed of 138- a nd 47-kDa polypeptides. Its isoelectric point is 6.8. This polymerase activity is strongly inhibited by N-ethylmaleimide, aphidicolin, and h igh KCl concentration but is relatively insensitive to 2',3' dideoxyth ymidine 5'-triphosphate. There was no reaction in an immunological tes t using monoclonal antibody against Drosophila DNA polymerase alpha. I n a final purification step, this polymerase activity was accompanied by 3' --> 5' exonuclease activity as expected proofreading activity. T his polymerase activity is remarkably stimulated by mouse proliferatin g cell nuclear antigen, which is structurally and immunologically very similar to a Drosophila counterpart. These properties clearly indicat e this enzyme belongs to the category of DNA polymerase delta.