title: RNA specificity and regulation of catalysis in the eukaryotic polynucleotide kinase Clp1.
abstract: RNA-specific polynucleotide kinases of the Clp1 subfamily are key components of various RNA maturation pathways. However, the structural basis explaining their substrate specificity and the enzymatic mechanism is elusive. Here, we report crystal structures of Clp1 from Caenorhabditis elegans (ceClp1) in a number of nucleotide- and RNA-bound states along the reaction pathway. The combined structural and biochemical analysis of ceClp1 elucidates the RNA specificity and lets us derive a general model for enzyme catalysis of RNA-specific polynucleotide kinases. We identified an RNA binding motif referred to as "clasp" as well as a conformational switch that involves the essential Walker A lysine (Lys127) and regulates the enzymatic activity of ceClp1. Structural comparison with other P loop proteins, such as kinases, adenosine triphosphatases (ATPases), and guanosine triphosphatases (GTPases), suggests that the observed conformational switch of the Walker A lysine is a broadly relevant mechanistic feature.
authors.affiliation: Department of Biomolecular Mechanisms, Max Planck Institute for Medical Research, Heidelberg 69120, Germany.
authors.forename: Aytac
authors.initials: A
authors.lastname: Dikfidan
authors.affiliation: Department of Biomolecular Mechanisms, Max Planck Institute for Medical Research, Heidelberg 69120, Germany; Institute for Chemistry and Biochemistry/Laboratory of Structural Biochemistry, Freie Universität Berlin, Berlin 14195, Germany.
authors.forename: Bernhard
authors.initials: B
authors.lastname: Loll
authors.affiliation: Department of Biomolecular Mechanisms, Max Planck Institute for Medical Research, Heidelberg 69120, Germany.
authors.forename: Cathleen
authors.initials: C
authors.lastname: Zeymer
authors.affiliation: Department of Biomolecular Mechanisms, Max Planck Institute for Medical Research, Heidelberg 69120, Germany.
authors.forename: Iris
authors.initials: I
authors.lastname: Magler
authors.affiliation: Research Institute of Molecular Pathology, Vienna 1030, Austria.
authors.forename: Tim
authors.initials: T
authors.lastname: Clausen
authors.affiliation: Department of Biomolecular Mechanisms, Max Planck Institute for Medical Research, Heidelberg 69120, Germany. Electronic address: anton.meinhart@mpimf-heidelberg.mpg.de.
authors.forename: Anton
authors.initials: A
authors.lastname: Meinhart
mesh_terms: D000251:Adenosine Triphosphatases / Q000648:ultrastructure; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D017173:Caenorhabditis elegans / Q000201:enzymology* / Q000235:genetics; D029742:Caenorhabditis elegans Proteins; D002384:Catalysis; D018360:Crystallography, X-Ray; D020558:GTP Phosphohydrolases / Q000648:ultrastructure; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000737:chemistry* / Q000235:genetics / Q000648:ultrastructure; D017434:Protein Structure, Tertiary; D012313:RNA / Q000096:biosynthesis; D011130:RNA Ligase (ATP) / Q000235:genetics / Q000378:metabolism / Q000648:ultrastructure*; D016601:RNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000648:ultrastructure; D013379:Substrate Specificity
chemical_list: D029742:Caenorhabditis elegans Proteins; D016601:RNA-Binding Proteins; D012313:RNA; D017853:Phosphotransferases (Alcohol Group Acceptor); C000591242:clpf-1 protein, C elegans; D000251:Adenosine Triphosphatases; D020558:GTP Phosphohydrolases; D011130:RNA Ligase (ATP)