Enhancing the prebiotic relevance of a set of covalently self-assembling, autorecombining RNAs through in vitro selection.
An in vitro form of the self-splicing group-I intron interrupting the Azoarcus tRNA(Ile) was shortened by ~10% with the removal of helix P6a. This deletion reduced the reverse-splicing activity of the ribozyme about 10-fold. Through in vitro selection, this activity was restored in several low-error mutants. A number of mutations were found that improved reverse-splicing activity through both increased k (obs) and better folding. The deletion mutant could be fragmented into as many as three discrete pieces, which, when incubated together, were capable of covalent self-assembly through energy-neutral transesterification reactions, a process called autorecombination. A subset of the mutations identified through in vitro selection for reverse-splicing were exaptations in that they were also shown to augment the autorecombination reactions, leading to higher yields of covalently self-assembled products, making this the smallest such system yet discovered.
Burton
Aaron S
AS
Department of Chemistry, Portland State University, OR 97207, USA.
Lehman
Niles
N
D020627:Azoarcus / Q000235:genetics; D001483:Base Sequence; D002384:Catalysis; D019020:Directed Molecular Evolution / Q000379:methods*; D004353:Drug Evaluation, Preclinical / Q000379:methods; D006860:Hydrogen Bonding; D066298:In Vitro Techniques; D008954:Models, Biological; D016297:Mutagenesis, Site-Directed; D009690:Nucleic Acid Conformation; D056692:Prebiotics / Q000032:analysis; D012329:RNA, Bacterial / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D016337:RNA, Catalytic / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification* / Q000378:metabolism; D045522:Recombinases / Q000235:genetics; D011995:Recombination, Genetic / Q000235:genetics*; D017384:Sequence Deletion
D056692:Prebiotics; D012329:RNA, Bacterial; D016337:RNA, Catalytic; D045522:Recombinases