The G-patch activators Pfa1 and PINX1 exhibit different modes of interaction with the Prp43 RNA helicase.

Prp43 is a DEAH-box RNA helicase involved in both splicing and ribosome biogenesis. Its activities are directly stimulated by several co-activators that share a G-patch domain. The substrates of Prp43, its mechanism of action and the modes of interaction with and activation by G-patch proteins have been only partially characterized. We investigated how Pfa1 and PINX1, two G-patch proteins involved in ribosome biogenesis, interact with Prp43. We demonstrate that a protruding loop connecting the β4 and β5 strands of Prp43 OB fold is crucial for the binding of the G-patch domain of Pfa1. However, neither this loop nor the entire OB fold of Prp43 is essential for PINX1 binding. We conclude that the binding modes of Pfa1 and PINX1 G-patches to Prp43 are different. Nevertheless, stimulation of the ATPase and helicase activities of Prp43 by both full-length Pfa1 and PINX1 requires the β4-β5 loop. Moreover, we show that disruption of this loop completely abrogates Prp43 activity during yeast ribosome biogenesis but does not prevent its integration within pre-ribosomal particles. We propose that the β4-β5 loop plays a crucial role in the transmission of conformational changes induced by binding of the G-patch to Prp43 active site and substrate RNA.