PURIFICATION AND CHARACTERIZATION OF AN ENDONUCLEASE FROM XENOPUS-LAEVIS OVARIES WHICH ACCURATELY PROCESSES THE 3' TERMINUS OF HUMAN PRE-TRANSFER RNA(I)MET (3' PRE-TRANSFER RNASE)

  • 1 January 1985
    • journal article
    • research article
    • Vol. 260  (15) , 9002-9008
Abstract
The primary transcript of the human .**GRAPHIC**. gene undergoes accurate processing to a mature 72-nucleotide species by activating present in the high speed supernatant of X. laevis ovarian extracts. The purification and characterization of the enzyme which processes the 3'' terminus of the human .**GRAPHIC**. species is described. The activity was purified .apprx. 500-fold from a high speed supernatant of X. laevis ovarian extracts by standard methods. It appears to function as a single polypeptide with a MW of .apprx. 97,400. The enzyme generates the mature 3'' terminus with a single endonucleolytic cut, also yielding the intact 3'' trailer. The endonuclease has a striking preference for the 5'' processed .**GRAPHIC**. exhibiting little or no activity in vitro on the intact primary transcript. The enzyme acts similarly with the pre-tRNAAla species of Bombyx mori, suggesting that it possesses a broad substrate range. The requirement of the 3'' processing endonuclease for a processed 5'' terminus suggests that eukaryotic pre-tRNA processing should follow an ordered cutting sequence in vivo with processing of the 5'' leader preceding 3'' end maturation.