METAL COORDINATION SITES THAT CONTRIBUTE TO STRUCTURE AND CATALYSIS IN THE GROUP-I INTRON FROM TETRAHYMENA

We have used nucleoside phosphorothioates (NTPalphaS) and a substituti on-interference method to identify phosphate oxygens that appear to be important to guanosine cofactor addition in the self-splicing group I intron from Tetrahymena thermophila. For the majority of these phosph ate oxygens, however, the effect of NTPalphaS substitution is signific antly reduced in reactions containing the added presence of manganese ion (Mn2+) relative to magnesium ion (Mg2+) alone. The observed ''resc ue'' of the NTPalphaS effect at these positions is thought to be due t o the larger affinity of Mn2+ for sulfur. These data suggest the direc t coordination of divalent metal ions within the highly conserved cata lytic core of the Tetrahymena intron. Because many of these metal bind ing sites appear to be in positions of close backbone-backbone approac h, and adjacent to the guanosine binding site the splice junction, we suggest roles for the corresponding ions in stabilizing tertiary struc ture and substrate recognition and as participants in catalysis.