Probing force-induced unfolding intermediates of a single staphylococcal nuclease molecule and the effect of ligand binding |
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Authors: | Ishii Takaaki Murayama Yoshihiro Katano Atsuto Maki Kosuke Kuwajima Kunihiro Sano Masaki |
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Institution: | a Department of Physics, Graduate School of Science, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan b Department of Applied Physics, Graduate School of Engineering, Tokyo University of Agriculture and Technology, 2-24-16 Naka-cho, Koganei-shi, Tokyo 184-8588, Japan c Graduate School of Science, Nagoya University, Fro-cho, Chikusa-ku, Nagoya City, Aichi 464-8602, Japan d Okazaki Institute for Integrative Bioscience, and Institute for Molecular Science, National Institutes of Natural Sciences, 5-1 Higashiyama, Myodaiji, Okazaki, Aichi 444-8787, Japan e Department of Functional Molecular Science, School of Physical Sciences, Graduate University for Advanced Studies (SOKENDAI), 5-1 Higashiyama, Myodaiji, Okazaki, Aichi 444-8787, Japan |
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Abstract: | Single-molecule manipulation techniques have given experimental access to unfolding intermediates of proteins that are inaccessible in conventional experiments. A detailed characterization of the intermediates is a challenging problem that provides new possibilities for directly probing the energy landscape of proteins. We investigated single-molecule mechanical unfolding of a small globular protein, staphylococcal nuclease (SNase), using atomic force microscopy. The unfolding trajectories of the protein displayed sub-molecular and stochastic behavior with typical lengths corresponding to the size of the unfolded substructures. Our results support the view that the single protein unfolds along multiple pathways as suggested in recent theoretical studies. Moreover, we found the drastic change, caused by the ligand and inhibitor bindings, in the mechanical unfolding dynamics. |
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Keywords: | Atomic force microscope (AFM) Staphylococcal nuclease (SNase) Mechanical unfolding Unfolding intermediate Ligand-binding complex |
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