Title

The Mechanism of HdeA Unfolding and Chaperone Activation

Authors

Loïc Salmon, Institute of Molecular Biology and Biophysics, ETH Zürich
Frederick Stull, Howard Hughes Medical Institute, Department of Molecular, Cellular, and Developmental Biology, University of Michigan, Department of Molecular, Cellular, and Developmental Biology, University of Michigan
Sabrina Sayle, Department of Molecular, Cellular, and Developmental Biology, University of Michigan
Claire Cato, Department of Biological Chemistry, University of Michigan Medical School
Şerife Akgül, Institute für Physikalische Biologie, Heinrich-Heine-Universität Düsseldorf
Linda Foit, Howard Hughes Medical Institute, Department of Molecular, Cellular, and Developmental Biology, University of Michigan, Department of Molecular, Cellular, and Developmental Biology, University of Michigan
Logan S. Ahlstrom, Howard Hughes Medical Institute, Department of Molecular, Cellular, and Developmental Biology, University of Michigan, Department of Molecular, Cellular, and Developmental Biology, University of Michigan
Elan Z. Eisenmesser, Department of Biochemistry and Molecular Genetics, University of Colorado Denver
Hashim M. Al-Hashimi, Department of Biochemistry, Duke University Medical Center, Department of Chemistry, Duke University
James C.A. Bardwell, Howard Hughes Medical Institute, Department of Molecular, Cellular, and Developmental Biology, University of Michigan, Department of Molecular, Cellular, and Developmental Biology, University of Michigan
Scott Horowitz, Department of Chemistry & Biochemistry and the Knoebel Institute for Healthy Aging, University of DenverFollow

Document Type

Article

Publication Date

11-11-2017

Keywords

Chaperone, Protein folding, Acid, NMR

Organizational Units

Knoebel Institute for Healthy Aging

Abstract

HdeA is a periplasmic chaperone that is rapidly activated upon shifting the pH to acidic conditions. This activation is thought to involve monomerization of HdeA. There is evidence that monomerization and partial unfolding allow the chaperone to bind to proteins denatured by low pH, thereby protecting them from aggregation. We analyzed the acid-induced unfolding of HdeA using NMR spectroscopy and fluorescence measurements, and obtained experimental evidence suggesting a complex mechanism in HdeA's acid-induced unfolding pathway, as previously postulated from molecular dynamics simulations. Counterintuitively, dissociation constant measurements show a stabilization of the HdeA dimer upon exposure to mildly acidic conditions. We provide experimental evidence that protonation of Glu37, a glutamate residue embedded in a hydrophobic pocket of HdeA, is important in controlling HdeA stabilization and thus the acid activation of this chaperone. Our data also reveal a sharp transition from folded dimer to unfolded monomer between pH 3 and pH 2, and suggest the existence of a low-populated, partially folded intermediate that could assist in chaperone activation or function. Overall, this study provides a detailed experimental investigation into the mechanism by which HdeA unfolds and activates.

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