Enhancing the prebiotic relevance of a set of covalently self-assembling, autorecombining RNAs through in vitro selection

J Mol Evol. 2010 Mar;70(3):233-41. doi: 10.1007/s00239-010-9325-3. Epub 2010 Mar 3.

Abstract

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.

Publication types

  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Azoarcus / genetics
  • Base Sequence
  • Catalysis
  • Directed Molecular Evolution / methods*
  • Drug Evaluation, Preclinical / methods
  • Hydrogen Bonding
  • In Vitro Techniques
  • Models, Biological
  • Mutagenesis, Site-Directed
  • Nucleic Acid Conformation
  • Prebiotics* / analysis
  • RNA, Bacterial / chemistry
  • RNA, Bacterial / genetics
  • RNA, Bacterial / isolation & purification
  • RNA, Bacterial / metabolism
  • RNA, Catalytic / chemistry
  • RNA, Catalytic / genetics*
  • RNA, Catalytic / isolation & purification*
  • RNA, Catalytic / metabolism
  • Recombinases / genetics
  • Recombination, Genetic / genetics*
  • Sequence Deletion

Substances

  • Prebiotics
  • RNA, Bacterial
  • RNA, Catalytic
  • Recombinases