An Overlapping Region between the Two Terminal Folding Units of the Outer Surface Protein A (OspA) Controls Its Folding Behavior. Issue 12 (8th June 2018)

• Record Type: Journal Article
• Authors: Makabe, Koki; Nakamura, Takashi; Dhar, Debanjan; Ikura, Teikichi; Koide, Shohei; Kuwajima, Kunihiro
• Is Part Of: Journal of molecular biology. Volume 430:Issue 12(2018)
• Page Start: 1799
• Page End: 1813
• Publication Date: 2018-06-08
• Subjects: OspA outer surface protein A -- SLB single-layer β-sheet; multi-domain protein -- beta-sheet -- on-pathway intermediate; Molecular biology -- Periodicals; Biology -- Periodicals; Biochemistry -- Periodicals; Bacteriology -- Periodicals; Molecular Biology -- Periodicals; Biochemistry -- Periodicals; Biologie moléculaire -- Périodiques; Biologie -- Périodiques; Biochimie -- Périodiques; Moleculaire biologie; Biochemistry; Biology; Molecular biology; Periodicals; 572.805
• DOI: 10.1016/j.jmb.2018.04.025
• Languages: English
• ISSNs: 0022-2836

Abstract

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. Graphical abstract: Highlights: Elucidation of mechanisms of multi-domain protein folding is an important issue. A two-domain protein OspA is a useful model for studying the multi-domain folding. We show evidence for the presence of an on-pathway folding intermediate in OspA. A rigid and stable intervening region between the domains played a critical role. The presence of such a region can lead to coupled folding behavior of the domains.