Prp43p contains a processive helicase structural architecture with a specific regulatory domain

Abstract

The DEAH/RNA helicase A (RHA) helicase family comprises proteins involved in splicing, ribosome biogenesis and transcription regulation. We report the structure of yeast Prp43p, a DEAH/RHA helicase remarkable in that it functions in both splicing and ribosome biogenesis. Prp43p displays a novel structural architecture with an unforeseen homology with the Ski2-like Hel308 DNA helicase. Together with the presence of a b-hairpin in the second RecA-like domain, Prp43p contains all the structural elements of a processive helicase. Moreover, our structure reveals that the C-terminal domain contains an oligonucleotide/oligosaccharide-binding (OB)-fold placed at the entrance of the putative nucleic acid cavity. Deletion or mutations of this domain decrease the affinity of Prp43p for RNA and severely reduce Prp43p ATPase activity in the presence of RNA. We also show that this domain constitutes the binding site for the G-patch-containing domain of Pfa1p. We propose that the C-terminal domain, specific to DEAH/RHA helicases, is a central player in the regulation of helicase activity by binding both RNA and G-patch domain proteins.

Structural basis for functional cooperation between tandem helicase cassettes in Brr2-mediated remodeling of the spliceosome

“…D and E). Therefore, S1087L abrogates functionally important S1087-RNA contacts and/or counteracts conformational changes in the ratchet helix, which have been suggested in related DEAH helicases (19). Because S1087L has no discernible effect on the folding of Brr2, it is likely that this and perhaps other RP33-linked Brr2 variants are incorporated into spliceosomes in vivo.…”

Functions and regulation of the Brr2 RNA helicase during splicing

“…In these enzymes, the N-terminal extensions seem to stabilize an inactive relative orientation of the 2 RecA-like domains, with the surfaces that have to come together for ATP and RNA binding remote from each other. In contrast, the N-terminal extensions of Prp43, 58,59,71 yeast Mtr4 68,69 and Neurospora crassa frequency-interacting RNA helicase (FRH) 78 together with accessory domains stabilize a relative orientation of the RecA-domains that resembles the productive conformation (Fig. 3D-F).…”

“…E.g., while the Ski2-like DNA helicase, Hel308, contains a HLH domain (which improves coupling of ATPase and unwinding activities 67 ) following the ratchet domain as also seen in Brr2, the Ski2-like RNA helicase, Mtr4, contains an arch-like domain inserted into its core (which aids in the recognition of specific substrates 68,69 and modulates helicase activity 70 ). The DEAH/RHA helicases Prp2, Prp16, Prp22 and Prp43, in contrast, contain a C-terminal OBfold, 58,59,71,72 which in Prp43 stimulates RNA binding and ATPase activity. 59 Together with Brr2, only very few other helicases, including yeast Slh1p 73 and human ASCC3, 74 exhibit 2 helicase cassettes.…”

“…The DEAH/RHA helicases Prp2, Prp16, Prp22 and Prp43, in contrast, contain a C-terminal OBfold, 58,59,71,72 which in Prp43 stimulates RNA binding and ATPase activity. 59 Together with Brr2, only very few other helicases, including yeast Slh1p 73 and human ASCC3, 74 exhibit 2 helicase cassettes. These enzymes form a small clade within the Ski2-like family.…”

“…In contrast, in DEAH/RHA helicases an element equivalent to the separator loop (termed "5 0 HP") has been suggested to mainly control access to the single-stranded RNA binding site. 58,59 Comparative modeling 41,51,52 and a recent electron cryomicroscopic (cryo-EM) structure of a yeast U4/U6 U5 trisnRNP 12 showed that Brr2 engages its U4/U6 di-snRNA substrate in a similar manner as Hel308. A single-stranded region of the U4 snRNA is threaded through the central tunnel of the NC between the RNA-binding motifs of the RecA and HB domains, with 3 0 -portions of the RNA extending toward the CC (Fig.…”