title: Xeroderma pigmentosum group C protein possesses a high affinity binding site to human centrin 2 and calmodulin.
abstract: Human centrin 2 (HsCen2), a member of the EF-hand superfamily of Ca2+-binding proteins, is commonly associated with centrosome-related structures. The protein is organized in two domains, each containing two EF-hand motifs, but only the C-terminal half exhibits Ca2+ sensor properties. A significant fraction of HsCen2 is localized in the nucleus, where it was recently found associated with the xeroderma pigmentosum group C protein (XPC), a component of the nuclear excision repair pathway. Analysis of the XPC sequence (940 residues), using a calmodulin target recognition software, enabled us to predict two putative binding sites. The binding properties of the two corresponding peptides were investigated by isothermal titration calorimetry. Only one of the peptides (P1-XPC) interacts strongly (Ka = 2.2 x 10(8) m-1, stoichiometry 1:1) with HsCen2 in a Ca2+-dependent manner. This peptide also binds, with a similar affinity (Ka = 1.1 x 10(8) m-1) to a C-terminal construct of HsCen2, indicating that the interaction with the integral protein is mainly the result of the contribution of the C-terminal half. The second peptide (P2-XPC) failed to show any detectable binding either to HsCen2 or to its C-terminal lobe. The two peptides interact with different affinities and mechanisms with calmodulin. Circular dichroism and nuclear magnetic resonance were used to structurally characterize the complex formed by the C-terminal domain of HsCen2 with P1-XPC.
authors.affiliation: INSERM U350 and Institut Curie-Recherche, Centre Universitaire, Batiments 110-112, 91405 Orsay, France.
authors.forename: Aurel
authors.initials: A
authors.lastname: Popescu
authors.forename: Simona
authors.initials: S
authors.lastname: Miron
authors.forename: Yves
authors.initials: Y
authors.lastname: Blouquit
authors.forename: Patricia
authors.initials: P
authors.lastname: Duchambon
authors.forename: Petya
authors.initials: P
authors.lastname: Christova
authors.forename: Constantin T
authors.initials: CT
authors.lastname: Craescu
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites / Q000235:genetics; D020013:Calcium Signaling; D002135:Calcium-Binding Proteins; D002147:Calmodulin / Q000378:metabolism*; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D046911:Macromolecular Substances; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: C472752:CETN2 protein, human; D002135:Calcium-Binding Proteins; D002147:Calmodulin; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D046911:Macromolecular Substances; D010446:Peptide Fragments; D011994:Recombinant Proteins; C086984:XPC protein, human