95771 20230921135433.0 doi 10.3390/biom10091313 sideral 120208 ART-2020-120208 eng Neira, J.L. A phosphorylation-induced switch in the nuclear localization sequence of the intrinsically disordered nupr1 hampers binding to importin 2020 Access copy available to the general public Unrestricted Several carrier proteins are involved in protein transport from the cytoplasm to the nucleus in eukaryotic cells. One of those is importin a, of which there are several human isoforms; among them, importin a3 (Impa3) has a high flexibility. The protein NUPR1, a nuclear protein involved in the cell-stress response and cell cycle regulation, is an intrinsically disordered protein (IDP) that has a nuclear localization sequence (NLS) to allow for nuclear translocation. NUPR1 does localize through the whole cell. In this work, we studied the affinity of the isolated wild-type NLS region (residues 54–74) of NUPR1 towards Impa3 and several mutants of the NLS region by using several biophysical techniques and molecular docking approaches. The NLS region of NUPR1 interacted with Impa3, opening the way to model the nuclear translocation of disordered proteins. All the isolated NLS peptides were disordered. They bound to Impa3 with low micromolar affinity (1.7–27 µM). Binding was hampered by removal of either Lys65 or Lys69 residues, indicating that positive charges were important; furthermore, binding decreased when Thr68 was phosphorylated. The peptide phosphorylated at Thr68, as well as four phospho-mimetic peptides (all containing the Thr68Glu mutation), showed the presence of a sequential NN(i, i + 1) nuclear Overhauser effect (NOE) in the 2D-1H-NMR (two-dimensional–proton NMR) spectra, indicating the presence of turn-like conformations. Thus, the phosphorylation of Thr68 modulates the binding of NUPR1 to Impa3 by a conformational, entropy-driven switch from a random-coil conformation to a turn-like structure. info:eu-repo/grantAgreement/ES/DGA/B25-17R info:eu-repo/grantAgreement/ES/DGA/E45-17R info:eu-repo/grantAgreement/ES/ISCIII/CPII13-00017 info:eu-repo/grantAgreement/ES/ISCIII-ERDF-ESF/PI15-00663-Investing in your future info:eu-repo/grantAgreement/ES/ISCIII-ERDF-ESF/PI18-00349-Investing in your future info:eu-repo/grantAgreement/ES/MCIU-AEI-FEDER/BFU2016-78232-P info:eu-repo/grantAgreement/ES/MCIU-AEI-FEDER/RTI2018-097991-B-I00 info:eu-repo/semantics/openAccess by http://creativecommons.org/licenses/by/3.0/es/ 4.879 2020 BIOCHEMISTRY & MOLECULAR BIOLOGY 96 / 296 = 0.324 2020 Q2 T1 1.125 2020 Molecular Biology 2020 Q2 Biochemistry 2020 Q2 info:eu-repo/semantics/article info:eu-repo/semantics/publishedVersion Rizzuti, B. Jiménez-Alesanco, A. Universidad de Zaragoza (orcid)0000-0003-4726-7821 Palomino-Schätzlein, M. Abián, O. Universidad de Zaragoza (orcid)0000-0001-5664-1729 Velázquez-Campoy, A. Universidad de Zaragoza (orcid)0000-0001-5702-4538 Iovanna, J.L. 1002 060 Universidad de Zaragoza Dpto. Bioq.Biolog.Mol. Celular Área Bioquímica y Biolog.Mole. 10, 9 (2020), 1313 [22 pp.] Biomolecules 2218-273X 637334 http://zaguan.unizar.es/record/95771/files/texto_completo.pdf Versión publicada 506640 http://zaguan.unizar.es/record/95771/files/texto_completo.jpg?subformat=icon icon Versión publicada oai:zaguan.unizar.es:95771 articulos driver 2023-09-21-13:30:34 ARTICLE