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Article

The Right-Handed Parallel β-Helix Topology of Erwinia chrysanthemi Pectin Methylesterase Is Intimately Associated with Both Sequential Folding and Resistance to High Pressure

1
Centre for Protein Engineering, Laboratory of Enzymology and Protein Folding, InBioS, University of Liège, Building B6C, Quartier Agora, Allée du 6 Août, 13, 4000 Liège, Sart-Tilman, Belgium
2
Department of Chemistry, University of Antwerp, Groenenborgerlaan 171, 2020 Antwerp, Belgium
*
Author to whom correspondence should be addressed.
This manuscript is dedicated to the memory of Christopher Martin Dobson (1949–2019), a leading British scientist in the field of protein folding and its link with misfolding diseases. He was an inspiring mentor to FM and AM during their postdoctoral years with him and had a major impact on their respective careers. The whole CMD community was dismayed by his death, and the world is a smaller and much sadder place without Chris. He will be remembered as an outstanding scientist but also as a great human being. He had genuine interest in how former members of his group were doing both professionally and also personally. We miss him a lot.
Academic Editor: Brigita Urbanc
Biomolecules 2021, 11(8), 1083; https://doi.org/10.3390/biom11081083
Received: 11 May 2021 / Revised: 1 July 2021 / Accepted: 3 July 2021 / Published: 22 July 2021
The complex topologies of large multi-domain globular proteins make the study of their folding and assembly particularly demanding. It is often characterized by complex kinetics and undesired side reactions, such as aggregation. The structural simplicity of tandem-repeat proteins, which are characterized by the repetition of a basic structural motif and are stabilized exclusively by sequentially localized contacts, has provided opportunities for dissecting their folding landscapes. In this study, we focus on the Erwinia chrysanthemi pectin methylesterase (342 residues), an all-β pectinolytic enzyme with a right-handed parallel β-helix structure. Chemicals and pressure were chosen as denaturants and a variety of optical techniques were used in conjunction with stopped-flow equipment to investigate the folding mechanism of the enzyme at 25 °C. Under equilibrium conditions, both chemical- and pressure-induced unfolding show two-state transitions, with average conformational stability (ΔG° = 35 ± 5 kJ·mol−1) but exceptionally high resistance to pressure (Pm = 800 ± 7 MPa). Stopped-flow kinetic experiments revealed a very rapid (τ < 1 ms) hydrophobic collapse accompanied by the formation of an extended secondary structure but did not reveal stable tertiary contacts. This is followed by three distinct cooperative phases and the significant population of two intermediate species. The kinetics followed by intrinsic fluorescence shows a lag phase, strongly indicating that these intermediates are productive species on a sequential folding pathway, for which we propose a plausible model. These combined data demonstrate that even a large repeat protein can fold in a highly cooperative manner. View Full-Text
Keywords: protein folding; parallel β-helix; repeat proteins; circular dichroism; high pressure; kinetic intermediate; sequential pathway; contact order; dry molten globule protein folding; parallel β-helix; repeat proteins; circular dichroism; high pressure; kinetic intermediate; sequential pathway; contact order; dry molten globule
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MDPI and ACS Style

Guillerm, J.; Frère, J.-M.; Meersman, F.; Matagne, A. The Right-Handed Parallel β-Helix Topology of Erwinia chrysanthemi Pectin Methylesterase Is Intimately Associated with Both Sequential Folding and Resistance to High Pressure. Biomolecules 2021, 11, 1083. https://doi.org/10.3390/biom11081083

AMA Style

Guillerm J, Frère J-M, Meersman F, Matagne A. The Right-Handed Parallel β-Helix Topology of Erwinia chrysanthemi Pectin Methylesterase Is Intimately Associated with Both Sequential Folding and Resistance to High Pressure. Biomolecules. 2021; 11(8):1083. https://doi.org/10.3390/biom11081083

Chicago/Turabian Style

Guillerm, Jessica, Jean-Marie Frère, Filip Meersman, and André Matagne. 2021. "The Right-Handed Parallel β-Helix Topology of Erwinia chrysanthemi Pectin Methylesterase Is Intimately Associated with Both Sequential Folding and Resistance to High Pressure" Biomolecules 11, no. 8: 1083. https://doi.org/10.3390/biom11081083

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