A 2'-PHOSPHOTRANSFERASE IMPLICATED IN TRANSFER-RNA SPLICING IS ESSENTIAL IN SACCHAROMYCES-CEREVISIAE

The last step of tRNA splicing in the yeast Saccharo myces cerevisiae is catalyzed by an NAD dependent 2'-phosphotransferase, which transfer s the splice junction 2'-phosphate from ligated tRNA to NAD to produce ADP-ribose 1 ''-2 '' cyclic phosphate. We have purified the phosphotr ansferase about 28,000-fold from yeast extracts and cloned its structu ral gene by reverse genetics. Expression of this gene (TPT1) in yeast or in Escherichia coli results in overproduction of 2'-phosphotransfer ase activity in extracts. Tpt1 protein is essential for vegetative gro wth in yeast, as demonstrated by gene disruption experiments. No obvio us binding motifs are found within the protein. Several candidate homo logs in other organisms are identified by searches of the data base, t he strongest of which is in Schizosaccharomyces pombe.