The structural elements involved in 5′ and 3′ splice site (SS) selection in a group II intron were analyzed. While 5′ SS selection appears to be defined by only one element, the EBS1-IBS1 pairing, four distinct structural components contribute to 3′ SS selection, one of which being analogous to the "internal guide sequence" described for group I introns. Moreover, some of the mutants analyzed during this study induce efficient 5′ SS hydrolysis and suggest how 5′ SS transesterification is selected against hydrolysis. Finally, the lariat structure was found to accelerate both steps of splicing, suggesting that it "locks" the ribozyme in an active configuration. © 1991.