A TRANSFER RNATRP INTRON ENDONUCLEASE FROM HALOBACTERIUM-VOLCANII - UNIQUE SUBSTRATE RECOGNITION PROPERTIES

  • 5 December 1988
    • journal article
    • research article
    • Vol. 263  (34) , 17951-17959
Abstract
We report on the properties of a partially purified tRNA intron endonuclease from the archaebacterium Halobacterium volcanii. This enzyme is capable of precise excision of the 104-nucleotide intron from halobacteria pre-tRNATrp substrates generated in vitro by T7 RNA polymerase transcription. The reaction requires divalent cations (Mg2+ or Ca2+) or spermidine, is inhibited by monovalent cations, and produces 5''-hydroxyl and 2'',3-cyclic phosphate termini. Unlike the universal substrate recognition properties characteristic of the eukaryotic tRNA intron endonucleases, this enzyme is specific for halophilic tRNATrp substrates. The partially purified enzyme is not capable of removing the intron from a yeast pre-tRNAPhe substrate. Analysis of the enzyme''s ability to cleave tRNATrp substrates lacking exon sequences demonstrated that the mature tRNA-like structure is not required in the substrate. A substrate retaining the intact intron and only the anticodon stem and loop exon regions was efficiently cleaved. Deletions within the intron indicated that the intron was not a primary site for recognition by the endonuclease; however, its presence affects the efficiency of the cleavage reaction. The possible relationship of this enzyme to other RNA endonucleases is discussed.