Novel RNA polymerization reaction catalyzed by a group I ribozyme.
We have converted a bacterial tRNA precursor containing a 205 nt self-splicing group I intron into a RNA enzyme that catalyzes polymerization of an external RNA substrate. The reaction involves transesterification steps analogous to both the forward and reverse exon ligation steps of group I splicing; as such it depends entirely on 3' splice site reactions. The RNA substrate is a 20 nt analogue of the ligated exons (E1.E2), whose 3' end resembles the 3' terminus of the intron RNA enzyme (IVS). The splice junction of the substrate is attacked by the 3' end of the intron, then the molecule displaces the original 3' terminal guanosine so that the new 3' terminus is brought into the active site and used as the attacking nucleophile in the next reaction. Polymerization occurs via a series of covalent enzyme-linked intermediates of the structure IVS.(E2)n, where n = 1 to > or = 18. The 5' exon accumulates during the course of the reaction and can attack the covalent intermediates to produce elongation products of structure E1.(E2)n, regenerating the intron RNA enzyme in unchanged form. In this manner, the enzyme converts 20 nt oligoribonucleotides into polyribonucleotides up to at least 180 nt by 10 nt increments. These results have significant implications for the evolution of RNA-based self-replicating systems.
Chowrira
B M
BM
Department of Microbiology and Molecular Genetics, Markey Center for Molecular Genetics, University of Vermont, Burlington 05405.
Berzal-Herranz
A
A
Burke
J M
JM
D000255:Adenosine Triphosphate / Q000378:metabolism; D001419:Bacteria / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D017931:DNA Primers; D005091:Exons; D007438:Introns; D046911:Macromolecular Substances; D008961:Models, Structural; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D010957:Plasmids; D016133:Polymerase Chain Reaction; D012322:RNA Precursors / Q000378:metabolism*; D012326:RNA Splicing; D012329:RNA, Bacterial / Q000378:metabolism*; D016337:RNA, Catalytic / Q000378:metabolism*; D012355:RNA, Transfer, Ile / Q000737:chemistry / Q000378:metabolism*; D013379:Substrate Specificity; D013698:Templates, Genetic
D017931:DNA Primers; D046911:Macromolecular Substances; D012322:RNA Precursors; D012329:RNA, Bacterial; D016337:RNA, Catalytic; D012355:RNA, Transfer, Ile; D000255:Adenosine Triphosphate