The deoxyribonuclease activity attributed to ribosome-inactivating proteins is due to contamination

The mode of action of ribosome-inactivating proteins (RIPs) has, For many y ears, been considered to be depurination of a specific adenyl residue of ri bosomal RNA, resulting in inhibition of protein synthesis. Recently, this v iew has been challenged by the observation that many RIP preparations have significant DNase activity in addition to their N-glycosidase activity. In this study, we have investigated the putative DNase activity of two RIPs, r icin and pokeweed antiviral protein (PAP), and show that, in both cases, th e DNase activity is due to the presence of contaminating nucleases. The N-g lycosidase and DNase activities of PAP were separately and specifically ina ctivated by chemical modification and heat. Gel filtration of ricin allowed physical separation of the two activities. Furthermore, neither recombinan t PAP nor recombinant ricin A-chain purified from Escherichia coli displaye d DNase activity.