Minimum ribonucleotide requirement for catalysis by the RNA hammerhead domain.
Several mixed DNA/RNA and 2'-O-methylribonucleotide/RNA analogues derived from the "hammerhead" domain of RNA catalysis have been prepared to study the minimum ribonucleotide requirement for catalytic activity. Oligodeoxyribonucleotides containing from seven to as few as four ribonucleotides are active in cleaving a substrate RNA. Predominantly deoxyribonucleotide-containing analogues have kcat values 20-300 and kcat/KM values approximately 100-2000 times lower than those of all-RNA ribozyme. In the case of predominantly 2'-O-methyl analogues, at least five ribonucleotides are needed to assure catalytic activity. In addition, both predominantly deoxyribonucleotide and 2'-O-methyl oligomers are at least 3 orders of magnitude more stable than an all-RNA ribozyme in incubations with RNase A and a yeast extract. These results suggest that the ribophosphate backbone is not a strict requirement for ribozyme-type catalysis. The identification of the four required ribonucleotides in the hammerhead catalytic domain provides valuable information for the rational design of chemical species having ribonuclease activities.
Yang
J H
JH
Département de Biochimie, Université de Montréal, Québec, Canada.
Usman
N
N
Chartrand
P
P
Cedergren
R
R
D001483:Base Sequence; D002384:Catalysis; D003854:Deoxyribonucleotides / Q000378:metabolism; D006868:Hydrolysis; D008745:Methylation; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D016337:RNA, Catalytic / Q000378:metabolism*; D012259:Ribonuclease, Pancreatic / Q000378:metabolism; D012265:Ribonucleotides / Q000378:metabolism*
D003854:Deoxyribonucleotides; D016337:RNA, Catalytic; D012265:Ribonucleotides; D012259:Ribonuclease, Pancreatic