Identification of a GUAAY Pentaloop Sequence Involved in a Novel RNA Loop-Helix Interaction

Large RNAs often utilize GNRA tetraloops as structural elements to stabilize the overall tertiary fold. These tetraloop-receptor (TR) interactions have a conserved geometry in which the tetraloop docks into the receptor at an angle of ~15° from the helix containing the receptor. Here, we show that the conserved GUAAY pentaloop found in domain III of group IIB1 introns participates in a novel class of RNA tertiary interaction with a geometry and mode of binding that are significantly different from that found in GNRA TR interactions. This pentaloop is highly conserved within the IIB1 class and interacts with the minor groove of the catalytic domain V. The base planes of the loop and receptor nucleotides are not coplanar and greatly deviate from standard A-minor motifs. The helical axis of the GUAAY stem loop diverges ~70° from the angle of insertion found in a typical GNRA TR interaction. Therefore, the loop architecture and insertion orientation are distinctive, with in vitro splicing data indicating that a GNRA tetraloop is incompatible at this position. The GUAAY pentaloop-receptor motif is also found in the structure of the eukaryotic thiamine pyrophosphate riboswitch in the context of a hexanucleotide loop sequence. We therefore propose, based on phylogenetic, structural, and biochemical data, that the GUAAY pentaloop-receptor interaction represents a novel structural motif that is present in multiple structured RNAs.