title: Identification of an RNA-dependent ATPase activity in mammalian U5 snRNPs.
abstract: Nuclear pre-mRNA splicing requires ATP at several steps from spliceosome assembly to product release. Here, we demonstrate that an integral component of the 20S U5 snRNP is an RNA-dependent ATPase. The ATPase activity of 20S U5 and 25S [U4/U6.U5] snRNPs purified by glycerol gradient centrifugation is strongly stimulated by homopolymeric RNA but not ssDNA. Purified 12S Ul and U2 snRNPs do not exhibit ATPase activity. Moreover, the U5-associated NTPase specifically hydrolyzes ATP and dATP. The additional purification of 20S U5 snRNPs by Mono Q chromatography does not affect the efficiency of ATP hydrolysis. Both U5 and tri-snRNPs bind ATP stoichiometrically in an RNA-independent manner. A candidate ATPase was identified by UV-irradiation of purified snRNPs with radiolabeled ATP. In the presence of homopolymeric RNA, the 200 kDa U5-specific protein is the major crosslinked protein, even in Mono Q-purified U5 snRNPs. The correlation between RNA-dependent ATPase activity in the U5 snRNP and the RNA-dependent onset of this crosslink strongly suggests that the 200 kDa protein is an RNA-dependent ATPase. Furthermore, both the formation of the crosslink and ATPase activity appear with a similar substrate specificity for ATP.
authors.affiliation: Institut für Molekularbiologie und Tumorforschung, Marburg, Germany.
authors.forename: B
authors.initials: B
authors.lastname: Laggerbauer
authors.forename: J
authors.initials: J
authors.lastname: Lauber
authors.forename: R
authors.initials: R
authors.lastname: Lührmann
mesh_terms: D000251:Adenosine Triphosphatases / Q000032:analysis* / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D006367:HeLa Cells; D006801:Humans; D008322:Mammals; D011869:Radioligand Assay; D017415:Ribonucleoprotein, U5 Small Nuclear / Q000032:analysis* / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D017415:Ribonucleoprotein, U5 Small Nuclear; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; C027900:RNA-dependent ATPase