A conserved element that stabilizes the group II intron active site

Abstract

The internal loop at the base of domain 3 (D3) is one of the most conserved and catalytically important elements of a group II intron. However, the location and molecular nature of its tertiary interaction partners has remained unknown. By employing a combination of site-directed photo-cross-linking and nucleotide analog interference suppression (NAIS), we show that the domain 3 internal loop (D3IL) interacts with the epsilon-epsilon' duplex, which is an active-site element located near the 5'-splice site in D1. Our data also suggest that the D3IL may interact with the bulge of D5, which is a critical active site component. The results of this and other recent studies indicate that the D3IL participates in a complex network of tertiary interactions involving epsilon-epsilon', the bulge of D5 and J23, and that it helps to optimize active site architecture by supporting interactions among these catalytic motifs. Our results are consistent with the role of D3 as a catalytic effector that enhances intron reactivity through active site stabilization.