A functional quantitative polymerase chain reaction assay for ricin, Shiga toxin, and related ribosome-inactivating proteins.
The potent toxins ricin, abrin, and other ribosome-inactivating proteins deadenylate a specific base in 28S ribosomal RNA that destroys ribosomes and leads to cell death. We have taken advantage of the fact that reverse transcriptase preferentially inserts an adenine opposite to an abasic site in RNA to create a quantitative polymerase chain reaction (PCR) assay to detect the damage. This assay detects as little as 30pg of ricin. We used the assay to study enzymatic properties of ricin such as pH and temperature optima (pH 4.5-5.0 and 60 degrees C).
Melchior
William B
WB
Division of Biochemical Toxicology, National Center for Toxicological Research, U.S. Food and Drug Administration, Jefferson, AR 72079, USA. william.melchior@fda.hhs.gov
Tolleson
William H
WH
D000036:Abrin / Q000378:metabolism; D000818:Animals; D017209:Apoptosis; D002460:Cell Line; D006863:Hydrogen-Ion Concentration; D016133:Polymerase Chain Reaction / Q000379:methods*; D012194:RNA-Directed DNA Polymerase / Q000378:metabolism; D051381:Rats; D054788:Ribosome Inactivating Proteins / Q000494:pharmacology*; D012276:Ricin / Q000494:pharmacology*; D022621:Shiga Toxin / Q000494:pharmacology*; D013696:Temperature
D000036:Abrin; D022621:Shiga Toxin; D012276:Ricin; D012194:RNA-Directed DNA Polymerase; D054788:Ribosome Inactivating Proteins