Purification of an Endonuclease from Adenovirus-Infected KB Cells

Abstract

The purification of an endonuclease from extracts of adenovirus-type-2 infected [human oral carcinoma] KB cells is described. Endonuclease activity can also be detected in extracts of uninfected KB cells, and the enzyme activities from extracts of uninfected and adenovirus-infected cells are very similar, if not identical. The enzyme has its maximal activity at pH 4.0. The enzyme found in uninfected and adenovirus-infected cells is strikingly different from an endonuclease isolated from calf serum. The endonuclease described is probably not a contaminant derived from the medium in which the KB cells were propagated. The endonuclease in crude extracts from uninfected or adenovirus-infected KB cells can be activated or its activity enhanced by treatment of the extracts with proteolytic enzymes, like pronase [EC 3.4.21.4 and EC 3.4.24.4] or trypsin [EC 3.4.21.4]. This activation is probably due to proteolytic cleavage of an inhibitor present in crude extracts of uninfected and adenovirus-type 2 infected KB cells. A 2nd endonuclease was found in extracts of infected and uninfected cells with optimal activity at pH 7.2, and this endonuclease can be separated from the one with a pH optimum at 4.0.