A conformationally restricted guanosine analog reveals the catalytic relevance of three structures of an RNA enzyme.
Recent studies indicate that RNA function can be enhanced by the incorporation of conformationally restricted nucleotides. Herein, we use 8-bromoguanosine, a nucleotide analog with an enforced syn conformation, to elucidate the catalytic relevance of ribozyme structures. We chose to study the lead-dependent ribozyme (leadzyme) because structural models derived from NMR, crystal, and computational (MC-Sym) studies differ in which of the three active site guanosines (G7, G9, or G24) have a syn glycosidic torsion angle. Kinetic assays were carried out on 8BrG variants at these three guanosine positions. These data indicate that an 8BrG24 leadzyme is hyperactive, while 8BrG7 and 8BrG9 leadzymes have reduced activity. These findings support the computational model of the leadzyme, rather than the NMR and crystal structures, as being the most relevant to phosphodiester bond cleavage.
Yajima
Rieko
R
Huck Institute for the Life Sciences, The Pennsylvania State University, University Park, PA 16802, USA.
Proctor
David J
DJ
Kierzek
Ryszard
R
Kierzek
Elzbieta
E
Bevilacqua
Philip C
PC
D001665:Binding Sites; D002384:Catalysis; D006151:Guanosine / Q000031:analogs & derivatives* / Q000737:chemistry; D007700:Kinetics; D008968:Molecular Conformation; D011487:Protein Conformation; D016337:RNA, Catalytic / Q000737:chemistry*
D016337:RNA, Catalytic; C113411:leadzyme; D006151:Guanosine; C035169:8-bromoguanosine