| Autor: |
Myrum C; Dr Einar Martens' Research Group for Biological Psychiatry, Center for Medical Genetics and Molecular Medicine, Haukeland University Hospital, 5009 Bergen, Norway., Baumann A; †Department of Biomedicine, University of Bergen, Jonas Lies vei 91, N-5009 Bergen, Norway., Bustad HJ; †Department of Biomedicine, University of Bergen, Jonas Lies vei 91, N-5009 Bergen, Norway., Flydal MI; †Department of Biomedicine, University of Bergen, Jonas Lies vei 91, N-5009 Bergen, Norway., Mariaule V; §European Molecular Biology Laboratory, Grenoble Outstation, 72 rue Jules Horowitz, BP181, 38042 Grenoble Cedex 9, France., Alvira S; ║Centro Nacional de Biotecnología (CNB-CSIC), Darwin 3, 28049 Madrid, Spain., Cuéllar J; ║Centro Nacional de Biotecnología (CNB-CSIC), Darwin 3, 28049 Madrid, Spain., Haavik J; †Department of Biomedicine, University of Bergen, Jonas Lies vei 91, N-5009 Bergen, Norway., Soulé J; †Department of Biomedicine, University of Bergen, Jonas Lies vei 91, N-5009 Bergen, Norway., Valpuesta JM; ║Centro Nacional de Biotecnología (CNB-CSIC), Darwin 3, 28049 Madrid, Spain., Márquez JA; §European Molecular Biology Laboratory, Grenoble Outstation, 72 rue Jules Horowitz, BP181, 38042 Grenoble Cedex 9, France., Martinez A; †Department of Biomedicine, University of Bergen, Jonas Lies vei 91, N-5009 Bergen, Norway., Bramham CR; †Department of Biomedicine, University of Bergen, Jonas Lies vei 91, N-5009 Bergen, Norway. |
| Abstrakt: |
The immediate early gene product Arc (activity-regulated cytoskeleton-associated protein) is posited as a master regulator of long-term synaptic plasticity and memory. However, the physicochemical and structural properties of Arc have not been elucidated. In the present study, we expressed and purified recombinant human Arc (hArc) and performed the first biochemical and biophysical analysis of hArc's structure and stability. Limited proteolysis assays and MS analysis indicate that hArc has two major domains on either side of a central more disordered linker region, consistent with in silico structure predictions. hArc's secondary structure was estimated using CD, and stability was analysed by CD-monitored thermal denaturation and differential scanning fluorimetry (DSF). Oligomerization states under different conditions were studied by dynamic light scattering (DLS) and visualized by AFM and EM. Biophysical analyses show that hArc is a modular protein with defined secondary structure and loose tertiary structure. hArc appears to be pyramid-shaped as a monomer and is capable of reversible self-association, forming large soluble oligomers. The N-terminal domain of hArc is highly basic, which may promote interaction with cytoskeletal structures or other polyanionic surfaces, whereas the C-terminal domain is acidic and stabilized by ionic conditions that promote oligomerization. Upon binding of presenilin-1 (PS1) peptide, hArc undergoes a large structural change. A non-synonymous genetic variant of hArc (V231G) showed properties similar to the wild-type (WT) protein. We conclude that hArc is a flexible multi-domain protein that exists in monomeric and oligomeric forms, compatible with a diverse, hub-like role in plasticity-related processes. |
