An inserted region of leucyl-tRNA synthetase plays a critical role in group I intron splicing.
Yeast mitochondrial leucyl-tRNA synthetase (LeuRS) binds to the bI4 intron and collaborates with the bI4 maturase to aid excision of the group I intron. Deletion analysis isolated the inserted LeuRS CP1 domain as a critical factor in the protein's splicing activity. Protein fragments comprised of just the LeuRS CP1 region rescued complementation of a yeast strain that expressed a splicing-defective LeuRS. Three-hybrid analysis determined that these CP1-containing LeuRS fragments, ranging from 214 to 375 amino acids, bound to the bI4 intron. In each case, interactions with only the LeuRS protein fragment specifically stimulated bI4 intron splicing activity. Substitution of a homologous CP1 domain from isoleucyl-tRNA synthetase or mutation within the LeuRS CP1 region of the smallest protein fragment abolished RNA binding and splicing activity. The CP1 domain is best known for its amino acid editing activity. However, these results suggest that elements within the LeuRS CP1 domain also play a novel role, independent of the full-length tRNA synthetase, in binding the bI4 group I intron and facilitating its self-splicing activity.
Rho
Seung Bae
SB
Department of Biology and Biochemistry, University of Houston, Houston, TX 77204-5001, USA.
Lincecum
Tommie L
TL
Martinis
Susan A
SA
D015153:Blotting, Western; D004926:Escherichia coli / Q000378:metabolism; D017353:Gene Deletion; D007438:Introns; D007935:Leucine-tRNA Ligase / Q000737:chemistry* / Q000235:genetics*; D008928:Mitochondria / Q000378:metabolism; D008957:Models, Genetic; D009154:Mutation; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012313:RNA / Q000378:metabolism; D012326:RNA Splicing; D020133:Reverse Transcriptase Polymerase Chain Reaction; D020798:Two-Hybrid System Techniques
D012313:RNA; D007935:Leucine-tRNA Ligase