Unnatural base pairs mediate the site-specific incorporation of an unnatural hydrophobic component into RNA transcripts.
Site-specific incorporation of a hydrophobic nucleotide analog into RNA, by T7 transcription mediated by unnatural base pairs, was developed. The nucleotide analog, 5-phenylethynyl-3-(beta-D-ribofuranosyl)pyridin-2-one 5-triphosphate (denoted by Ph-yTP), was chemically synthesized and then site-specifically incorporated by T7 RNA polymerase into RNA opposite the pairing partner, 2-amino-6-(2-thienyl)purine (denoted by s) in DNA templates. The introduction of Ph-y into a theophylline-binding RNA aptamer, in which a uridine in the internal loop was replaced by Ph-y, raised the thermal stability of the aptamer. Thus, this unnatural nucleotide analog would be useful for stabilizing RNA tertiary structures and complexes between RNA and other molecules.
Endo
Masayuki
M
Protein Synthesis Technology Team, Protein Research Group, RIKEN Genomic Sciences Center, 1-7-22 Suehiro-cho, Tsurumi-ku, Yokohama, Kanagawa 230-0045, Japan.
Mitsui
Tsuneo
T
Okuni
Taeko
T
Kimoto
Michiko
M
Hirao
Ichiro
I
Yokoyama
Shigeyuki
S
D020029:Base Pairing; D001665:Binding Sites; D012321:DNA-Directed RNA Polymerases; D057927:Hydrophobic and Hydrophilic Interactions; D009690:Nucleic Acid Conformation; D009711:Nucleotides / Q000138:chemical synthesis; D011687:Purines / Q000138:chemical synthesis*; D011725:Pyridines / Q000138:chemical synthesis*; D020871:RNA Stability; D012333:RNA, Messenger / Q000138:chemical synthesis* / Q000737:chemistry; D013806:Theophylline / Q000378:metabolism; D014158:Transcription, Genetic; D014764:Viral Proteins
D009711:Nucleotides; D011687:Purines; D011725:Pyridines; D012333:RNA, Messenger; D014764:Viral Proteins; D013806:Theophylline; C062210:bacteriophage T7 RNA polymerase; D012321:DNA-Directed RNA Polymerases