The deoxyribonuclease activity attributed to ribosome-inactivating proteins is due to contamination.
The mode of action of ribosome-inactivating proteins (RIPs) has, for many years, been considered to be depurination of a specific adenyl residue of ribosomal RNA, resulting in inhibition of protein synthesis. Recently, this view 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, ricin and pokeweed antiviral protein (PAP), and show that, in both cases, the DNase activity is due to the presence of contaminating nucleases. The N-glycosidase and DNase activities of PAP were separately and specifically inactivated by chemical modification and heat. Gel filtration of ricin allowed physical separation of the two activities. Furthermore, neither recombinant PAP nor recombinant ricin A-chain purified from Escherichia coli displayed DNase activity.
Day
P J
PJ
Department of Biological Sciences, University of Warwick, Coventry, UK.
Lord
J M
JM
Roberts
L M
LM
D002850:Chromatography, Gel; D004278:DNA, Superhelical / Q000235:genetics; D003851:Deoxyribonucleases / Q000302:isolation & purification / Q000378:metabolism*; D004047:Diethyl Pyrocarbonate / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D008274:Magnesium / Q000494:pharmacology; D009699:N-Glycosyl Hydrolases / Q000378:metabolism; D009690:Nucleic Acid Conformation; D010940:Plant Proteins / Q000302:isolation & purification / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics; D011994:Recombinant Proteins / Q000378:metabolism; D054788:Ribosome Inactivating Proteins; D054789:Ribosome Inactivating Proteins, Type 1; D054790:Ribosome Inactivating Proteins, Type 2; D012270:Ribosomes / Q000378:metabolism*; D012276:Ricin / Q000378:metabolism; D013696:Temperature
D004278:DNA, Superhelical; D004791:Enzyme Inhibitors; D010940:Plant Proteins; D012333:RNA, Messenger; D011994:Recombinant Proteins; D054789:Ribosome Inactivating Proteins, Type 1; D054790:Ribosome Inactivating Proteins, Type 2; D012276:Ricin; D003851:Deoxyribonucleases; D009699:N-Glycosyl Hydrolases; D054788:Ribosome Inactivating Proteins; C024275:pokeweed antiviral protein; D008274:Magnesium; D004047:Diethyl Pyrocarbonate