The genomic HDV ribozyme utilizes a previously unnoticed U-turn motif to accomplish fast site-specific catalysis

Abstract

The genome of the human hepatitis delta virus (HDV) harbors a self-cleaving catalytic RNA motif, the genomic HDV ribozyme, whose crystal structure shows the dangling nucleotides 5′ of the cleavage site projecting away from the catalytic core. This 5′-sequence contains a clinically conserved U − 1 that we find to be essential for fast cleavage, as the order of activity follows U − 1 > C − 1 > A − 1 > G − 1, with a >25-fold activity loss from U − 1 to G − 1. Terbium(III) footprinting detects conformations for the P1.1 stem, the cleavage site wobble pair and the A-minor motif of the catalytic trefoil turn that depend on the identity of the N − 1 base. The most tightly folded catalytic core, resembling that of the reaction product, is found in the U − 1 wild-type precursor. Molecular dynamics simulations demonstrate that a U − 1 forms the most robust kink around the scissile phosphate, exposing it to the catalytic C75 in a previously unnoticed U-turn motif found also, for example, in the hammerhead ribozyme and tRNAs. Strikingly, we find that the common structural U-turn motif serves distinct functions in the HDV and hammerhead ribozymes.