The effect of cytidine on the structure and function of an RNA ligase ribozyme.
A cytidine-free ribozyme with RNA ligase activity was obtained by in vitro evolution, starting from a pool of random-sequence RNAs that contained only guanosine, adenosine, and uridine. This ribozyme contains 74 nt and catalyzes formation of a 3',5'-phosphodiester linkage with a catalytic rate of 0.016 min(-1). The RNA adopts a simple secondary structure based on a three-way junction motif, with ligation occurring at the end of a stem region located several nucleotides away from the junction. Cytidine was introduced to the cytidine-free ribozyme in a combinatorial fashion and additional rounds of in vitro evolution were carried out to allow the molecule to adapt to this added component. The resulting cytidine-containing ribozyme formed a 3',5' linkage with a catalytic rate of 0.32 min(-1). The improved rate of the cytidine-containing ribozyme was the result of 12 mutations, including seven added cytidines, that remodeled the internal bulge loops located adjacent to the three-way junction and stabilized the peripheral stem regions.
Rogers
J
J
Department of Chemistry, The Scripps Research Institute, La Jolla, California 92037, USA.
Joyce
G F
GF
D001483:Base Sequence; D003562:Cytidine; D019020:Directed Molecular Evolution; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D011118:Polynucleotide Ligases; D016337:RNA, Catalytic / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
D016337:RNA, Catalytic; D003562:Cytidine; D011118:Polynucleotide Ligases