Uncompetitive inhibition by adenine of the RNA-N-glycosidase activity of ribosome-inactivating proteins.
Ricin is a member of the ribosome-inactivating protein (RIP) family with RNA-N-glycosidase activity which inactivates eukaryotic ribosomes by specifically removing adenine from the first adenosine of a highly conserved GAGA loop present in 28S rRNA. Free adenine protects ribosomes in cell-free systems from inactivation by ricin. Protection by adenine is highly specific, since AMP, adenosine and modified adenines (1-methyladenine and ethenoadenine) were completely ineffective. Kinetic analysis of the behaviour of adenine as inhibitor of the RNA-N-glycosidase reaction catalysed by ricin, Shiga-like toxin I and momordin, two other members of the RIP family, established that inhibition was of the uncompetitive type, the inhibitor binding to the enzyme-substrate complex. Adenine did not protect ribosomes from alpha-sarcin, an RNAase that inactivates ribosomes by cleaving the phosphodiester bond located in the GAGA loop at one nucleotide distance from the adenosine depurinated by the RNA-N-glycosidases. Adenine at the concentration of 1 mM lowered 1.5-fold the toxicity of ricin and 3.7-fold that of Shiga-like toxin I on Vero cells in culture. The same concentration of adenine decreased 2.4-fold the inactivation of isolated ribosomes by ricin, 2.8-fold the inactivation by Shiga-like toxin I and 20-fold that by momordin.
Pallanca
A
A
Dipartimento di Patologia Sperimentale dell'Università degli Studi di Bologna, Italy.
Mazzaracchio
R
R
Brigotti
M
M
Carnicelli
D
D
Alvergna
P
P
Sperti
S
S
Montanaro
L
L
D000225:Adenine / Q000494:pharmacology*; D000818:Animals; D001427:Bacterial Toxins / Q000494:pharmacology; D002522:Chlorocebus aethiops; D009699:N-Glycosyl Hydrolases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D054788:Ribosome Inactivating Proteins; D012270:Ribosomes / Q000187:drug effects*; D012276:Ricin / Q000494:pharmacology; D022622:Shiga Toxin 1; D014709:Vero Cells / Q000187:drug effects / Q000201:enzymology
D001427:Bacterial Toxins; D022622:Shiga Toxin 1; D012276:Ricin; D009699:N-Glycosyl Hydrolases; D054788:Ribosome Inactivating Proteins; D000225:Adenine