Abstract
Noncoding intron sequences present in precursor mRNAs need to be removed prior to translation, and they are excised via the spliceosome, a multimegadalton molecular machine composed of numerous protein and RNA components. The DEAH-box ATPase Prp2 plays a crucial role during pre-mRNA splicing as it ensures the catalytic activation of the spliceosome. Despite high structural similarity to other spliceosomal DEAH-box helicases, Prp2 does not seem to function as an RNA helicase, but rather as an RNA-dependent ribonucleoprotein particle-modifying ATPase. Recent crystal structures of the spliceosomal DEAH-box ATPases Prp43 and Prp22, as well as of the related RNA helicase MLE, in complex with RNA have contributed to a better understanding of how RNA binding and processivity might be achieved in this helicase family. In order to shed light onto the divergent manner of function of Prp2, an N-terminally truncated construct of Chaetomium thermophilum Prp2 was crystallized in the presence of ADP-BeF3- and a poly-U12 RNA. The refined structure revealed a virtually identical conformation of the helicase core compared with the ADP-BeF3-- and RNA-bound structure of Prp43, and only a minor shift of the C-terminal domains. However, Prp2 and Prp43 differ in the hook-loop and a loop of the helix-bundle domain, which interacts with the hook-loop and evokes a different RNA conformation immediately after the 3' stack. On replacing these loop residues in Prp43 by the Prp2 sequence, the unwinding activity of Prp43 was abolished. Furthermore, a putative exit tunnel for the γ-phosphate after ATP hydrolysis could be identified in one of the Prp2 structures.
Highlights
In eukaryotes, most precursor messenger RNAs contain noncoding intron sequences which need to be removed in order to obtain a mature mRNA that can serve as a template for translation
Noncoding intron sequences present in precursor mRNAs need to be removed prior to translation, and they are excised via the spliceosome, a multimegadalton molecular machine composed of numerous protein and RNA components
The ssRNA binds to an RNA-binding tunnel between the helicase core and the C-terminal domains, as previously reported for the spliceosomal DEAH-box ATPases Prp43 and Prp22 (Fig. 1a; Tauchert et al, 2017; Hamann et al, 2019; He et al, 2017)
Summary
Most precursor messenger RNAs (pre-mRNAs) contain noncoding intron sequences which need to be removed in order to obtain a mature mRNA that can serve as a template for translation. The vast majority of these intervening sequences are removed with the help of the spliceosome (Will & Luhrmann, 2011; Wahl et al, 2009; Matera & Wang, 2014). For each intron to be removed, the complex is formed de novo on a pre-mRNA. Since it has no preformed active site, compositional as well as conformational rearrangements ensure the formation of a catalytically active complex. Once the intron has been excised via two subsequent transesterification reactions, the complex is completely disassembled and each component is available for a new round of splicing
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