Dynamics of the intrinsically disordered protein NUPR1 in isolation and in its fuzzy complexes with DNA and prothymosin α |
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| Institution: | 1. Department of Biochemistry and Molecular Biophysics, Washington University School of Medicine, 660 South Euclid Avenue, St. Louis, MO 63110, USA;2. Department of Biomedical Engineering and Center for Biological Systems Engineering, Washington University in St. Louis, 1 Brookings Drive, St. Louis, MO 63130, USA;3. Department of Chemistry and Biochemistry, Utah State University, 0300 Old Main Hill, Logan, UT 84322, USA;1. Dept. Life and Environmental Sciences, Marche Polytechnic University, Ancona 60131, Italy;2. Biophysics Institute, National Research Council, Palermo 90143, Italy;3. Graz University of Technology, 8010 Graz, Austria;1. Bio21 Molecular Science and Biotechnology Institute, The University of Melbourne, Victoria 3010, Australia;2. The Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria 3052, Australia;3. Department of Medical Biology, The University of Melbourne, Victoria 3010, Australia;4. Olivia Newton-John Cancer Research Institute, Heidelberg, Victoria 3084, Australia;5. School of Cancer Medicine, La Trobe University, Melbourne, Victoria 3086, Australia;6. Department of Chemistry and Physics, La Trobe Institute for Molecular Science, Melbourne, Victoria 3086, Australia;1. Christian Doppler Laboratory for High-Content Structural Biology and Biotechnology, Department of Computational and Structural Biology, Max F. Perutz Laboratories, University of Vienna, Dr-Bohr-Gasse 9, A-1030 Vienna, Austria;2. Department of Computational and Structural Biology, Max F. Perutz Laboratories, University of Vienna, Campus Vienna Biocenter 5, A-1030 Vienna, Austria;3. Laboratoire des biomolécules, LBM, Département de chimie, École normale supérieure, PSL University, Sorbonne Université, CNRS, 75005 Paris, France;1. School of Bioscience, Indian Institute of Technology Kharagpur, Kharagpur, West Bengal 721302, India;2. Department of Chemical Engineering, Indian Institute of Technology Kharagpur, Kharagpur, West Bengal 721302, India;3. School of Energy Science and Engineering, Indian Institute of Technology Kharagpur, Kharagpur, West Bengal 721302, India |
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| Abstract: | Intrinsically disordered proteins (IDPs) explore diverse conformations in their free states and, a few of them, also in their molecular complexes. This functional plasticity is essential for the function of IDPs, although their dynamics in both free and bound states is poorly understood. NUPR1 is a protumoral multifunctional IDP, activated during the acute phases of pancreatitis. It interacts with DNA and other IDPs, such as prothymosin α (ProTα), with dissociation constants of ~0.5 μM, and a 1:1 stoichiometry. We studied the structure and picosecond-to-nanosecond (ps-ns) dynamics by using both NMR and SAXS in: (i) isolated NUPR1; (ii) the NUPR1/ProTα complex; and (iii) the NUPR1/double stranded (ds) GGGCGCGCCC complex. Our SAXS findings show that NUPR1 remained disordered when bound to either partner, adopting a worm-like conformation; the fuzziness of bound NUPR1 was also pinpointed by NMR. Residues with the largest values of the relaxation rates (R1, R1ρ, R2 and ηxy), in the free and bound species, were mainly clustered around the 30s region of the sequence, which agree with one of the protein hot-spots already identified by site-directed mutagenesis. Not only residues in this region had larger relaxation rates, but they also moved slower than the rest of the molecule, as indicated by the reduced spectral density approach (RSDA). Upon binding, the energy landscape of NUPR1 was not funneled down to a specific, well-folded conformation, but rather its backbone flexibility was kept, with distinct motions occurring at the hot-spot region. |
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