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J Mol Biol. 2018 Jun 8;430(12):1799-1813. doi: 10.1016/j.jmb.2018.04.025. Epub 2018 Apr 27.

An Overlapping Region between the Two Terminal Folding Units of the Outer Surface Protein A (OspA) Controls Its Folding Behavior.

Author information

1
Graduate School of Science and Engineering, Yamagata University, Jyonan 4-3-16, Yonezawa, Yamagata 992-8510, Japan; Okazaki Institute for Integrative Bioscience and Institute for Molecular Science, National Institutes of Natural Sciences, 5-1 Higashiyama, Myodaiji, Okazaki 444-8787, Japan; Department of Functional Molecular Science, SOKENDAI (The Graduate University for Advanced Studies), 5-1 Higashiyama, Myodaiji, Okazaki 444-8787, Japan. Electronic address: makabe@yz.yamagata-u.ac.jp.
2
Okazaki Institute for Integrative Bioscience and Institute for Molecular Science, National Institutes of Natural Sciences, 5-1 Higashiyama, Myodaiji, Okazaki 444-8787, Japan.
3
Laboratory of Structural Biology, School of Biomedical Science, Tokyo Medical and Dental University, Tokyo 113-8510, Japan.
4
Department of Biochemistry and Molecular Pharmacology, New York University School of Medicine, and Perlmutter Cancer Center at NYU Langone Health, New York, NY 10016, USA.
5
Okazaki Institute for Integrative Bioscience and Institute for Molecular Science, National Institutes of Natural Sciences, 5-1 Higashiyama, Myodaiji, Okazaki 444-8787, Japan; Department of Functional Molecular Science, SOKENDAI (The Graduate University for Advanced Studies), 5-1 Higashiyama, Myodaiji, Okazaki 444-8787, Japan; Department of Physics, School of Science, University of Tokyo, Bunkyo-ku, Tokyo 113-0033, Japan; School of Computational Sciences, Korea Institute for Advanced Study (KIAS), Dongdaemun-gu, Seoul 130-722, Korea.

Abstract

Although many naturally occurring proteins consist of multiple domains, most studies on protein folding to date deal with single-domain proteins or isolated domains of multi-domain proteins. Studies of multi-domain protein folding are required for further advancing our understanding of protein folding mechanisms. Borrelia outer surface protein A (OspA) is a β-rich two-domain protein, in which two globular domains are connected by a rigid and stable single-layer β-sheet. Thus, OspA is particularly suited as a model system for studying the interplays of domains in protein folding. Here, we studied the equilibria and kinetics of the urea-induced folding-unfolding reactions of OspA probed with tryptophan fluorescence and ultraviolet circular dichroism. Global analysis of the experimental data revealed compelling lines of evidence for accumulation of an on-pathway intermediate during kinetic refolding and for the identity between the kinetic intermediate and a previously described equilibrium unfolding intermediate. The results suggest that the intermediate has the fully native structure in the N-terminal domain and the single layer β-sheet, with the C-terminal domain still unfolded. The observation of the productive on-pathway folding intermediate clearly indicates substantial interactions between the two domains mediated by the single-layer β-sheet. We propose that a rigid and stable intervening region between two domains creates an overlap between two folding units and can energetically couple their folding reactions.

KEYWORDS:

beta-sheet; multi-domain protein; on-pathway intermediate

PMID:
29709572
DOI:
10.1016/j.jmb.2018.04.025

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