title: Synergic role of nucleophosmin three-helix bundle and a flanking unstructured tail in the interaction with G-quadruplex DNA.
abstract: Nucleophosmin (NPM1) is a nucleocytoplasmic shuttling protein, mainly localized at nucleoli, that plays a number of functions in ribosome biogenesis and export, cell cycle control, and response to stress stimuli. NPM1 is the most frequently mutated gene in acute myeloid leukemia; mutations map to the C-terminal domain of the protein and cause its denaturation and aberrant cytoplasmic translocation. NPM1 C-terminal domain binds G-quadruplex regions at ribosomal DNA and at gene promoters, including the well characterized sequence from the nuclease-hypersensitive element III region of the c-MYC promoter. These activities are lost by the leukemic variant. Here we analyze the NPM1/G-quadruplex interaction, focusing on residues belonging to both the NPM1 terminal three-helix bundle and a lysine-rich unstructured tail, which has been shown to be necessary for high affinity recognition. We performed extended site-directed mutagenesis and measured binding rate constants through surface plasmon resonance analysis. These data, supported by molecular dynamics simulations, suggest that the unstructured tail plays a double role in the reaction mechanism. On the one hand, it facilitates the formation of an encounter complex through long range electrostatic interactions; on the other hand, it directly contacts the G-quadruplex scaffold through multiple and transient electrostatic interactions, significantly enlarging the contact surface.
authors: From the Istituto di Biochimica e Biochimica Clinica, Università Cattolica del Sacro Cuore, 00168 Rome, Italy.
authors: Alessandro
authors: A
authors: Arcovito
authors: Ce.S.I. Centro Scienze dell'Invecchiamento, "Fondazione Università D'Annunzio," 66013 Chieti, Italy, Dipartimento di Scienze Biochimiche, Sapienza Università di Roma, 00185 Rome, Italy.
authors: Sara
authors: S
authors: Chiarella
authors: Dipartimento di Medicina Clinica, Sanità Pubblica, Scienze della Vita e dell'Ambiente, Università dell'Aquila, 67100 Coppito (L'Aquila), Italy.
authors: Stefano
authors: S
authors: Della Longa
authors: Istituto di Biologia, Medicina Molecolare e NanoBiotecnologie, Consiglio Nazionale delle Ricerche, 00185 Rome, Italy, and.
authors: Adele
authors: A
authors: Di Matteo
authors: Dipartimento di Scienze Biochimiche, Sapienza Università di Roma, 00185 Rome, Italy.
authors: Carlo
authors: C
authors: Lo Sterzo
authors: From the Istituto di Biochimica e Biochimica Clinica, Università Cattolica del Sacro Cuore, 00168 Rome, Italy.
authors: Giovanni Luca
authors: GL
authors: Scaglione
authors: Ce.S.I. Centro Scienze dell'Invecchiamento, "Fondazione Università D'Annunzio," 66013 Chieti, Italy, Dipartimento di Scienze Sperimentali e Cliniche, Università di Chieti "G. D'Annunzio," 66013 Chieti, Italy lfederici@unich.it.
authors: Luca
authors: L
authors: Federici
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D017931:DNA Primers; D054856:G-Quadruplexes; D056004:Molecular Dynamics Simulation; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D000090243:Nucleophosmin; D011485:Protein Binding; D017386:Sequence Homology, Amino Acid; D020349:Surface Plasmon Resonance
chemical_list: D017931:DNA Primers; D009687:Nuclear Proteins; D000090243:Nucleophosmin
keywords: Encounter Complex; Flanking Fuzziness; Intrinsically Disordered Protein; Leukemia; Molecular Dynamics; Protein-DNA Interaction; Surface Plasmon Resonance (SPR)