A novel group I intron-encoded endonuclease specific for the anticodon region of tRNA(fMet) genes.
Open reading frames (ORFs) are frequently inserted into group I self-splicing introns. These ORFs encode either maturases that are required for splicing of the intron or DNA endonucleases that promote intron mobility. A self-splicing intron in the tRNA(fMet) gene of Synechocystis PCC 6803, which has been proposed to have moved laterally within the cyanobacteria, contains an ORF that is unrelated to known intron-encoded endonucleases or maturases. Here, using an in vitro transcription-translation system, we show that this intronic ORF encodes a double-strand DNA endonuclease, I-Ssp6803I. I-Ssp6803I cleaves each strand of the intronless tRNA(fMet) gene adjacent to the anticodon triplet leaving 3 bp 3' extensions and has no activity at intron-exon boundaries. Using an in vitro cleavage assay and scanning deletion mutants of the intronless target site, the minimal recognition site was determined to be a partially palindromic 20 bp region encompassing the entire anticodon stem and loop of the tRNA(fMet) gene. I-Ssp6803I represents a novel intron-encoded DNA endonuclease and is the first example of a chromosomally encoded group I intron endonuclease in bacteria.
Department of Biological Sciences and Center for Molecular Genetics, University at Albany, State University of New York, 12222, USA.
R P
RP
Bonocora
D A
DA
Shub
D000595:Amino Acid Sequence; D000926:Anticodon / Q000235:genetics; D001483:Base Sequence; D000458:Cyanobacteria / Q000235:genetics*; D003850:Deoxyribonuclease I / Q000235:genetics* / Q000378:metabolism*; D007438:Introns / Q000235:genetics*; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D016366:Open Reading Frames / Q000235:genetics; D012358:RNA, Transfer, Met / Q000235:genetics*
D000926:Anticodon; D012358:RNA, Transfer, Met; D003850:Deoxyribonuclease I