The DNA polymerase in crude extracts of Drosophila melanogaster embryos sedimented at 9.0, 7.3, and 5.5 S on glycerol velocity gradients. The relative proportions of these enzymes depended on the method used to prepare the extract. Extracts of whole embryos contained the 7.3S and the 5.5S DNA polymerases and extracts of dechorionated embryos contained the 9.0S and 7.3S DNA polymerases. The porportion of the 5.5S DNA polymerase increased relative to the 7.3S DNA polymerase during storage of the extract of whole embryos. The protease inhibitor, phenylmethanesulfonyl fluoride, inhibited the formation of the 5.5S DNA polymerase, suggesting that it was proteolytically produced from the 7.3S DNA polymerase. This was demonstrated directly by converting the 7.3S DNA polymerase to the 5.5S DNA polymerase by treatment in vitro with trypsin. The degradation of the enzyme occurred without significant loss of DNA polymerase activity. It is further demonstrated that endogenous proteolysis reduced the chromatographic heterogeneity of the Drosophila DNA polymerase on diethylaminoethyl-Sephadex. When endogenous proteolysis was reduced, three forms of DNA polymerase were isolated by diethylaminoethylcellulose chromatography; two of these enzymes sedimented at 7.3S and the third sedimented at 9.0S. These results demonstrate the physical heterogeneity of the Drosophila DNA polymerase and suggest its similarity to vertebrate DNA polymerase-alpha.