Penning, Stephanie, Mitchell, Lachlan, Hong, Yaoqin, Cunliffe, Taylor, Subedi, Pramod, Wang, Geqing, Hor, Lilian, Totsika, Makrina, Paxman, Jason J., and Heras, Begoña (2025) Structural and functional specialization of Bordetella pertussis DsbA for pertussis toxin folding. Protein Science, 35 (1). 70421.

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Abstract

Disulphide bonds (Dsbs) are essential for the folding, stability, and function of many secreted and membrane-associated proteins in bacteria. In Gram-negative species, these bonds are introduced by the Dsb enzyme family, with DsbA acting as the primary thiol oxidase. While DsbA proteins share a conserved thioredoxin (TRX)-like fold, emerging evidence highlights substantial structural and functional divergence among pathogenic homologues. Here, we present the high-resolution crystal structure and functional characterization of BperDsbA, a DsbA homologue from Bordetella pertussis, the causative agent of whooping cough. BperDsbA adopts a canonical TRX fold with a CPHC active site and a threonine-containing cis-proline loop, but displays striking deviations from prototypical DsbAs. Notably, it contains a highly destabilizing catalytic Dsb, resulting in one of the most oxidizing redox potentials recorded for a DsbA enzyme. Surface electrostatic analysis reveals an unusual distribution of positive and negative charge around the active site, in contrast to the broadly hydrophobic catalytic surfaces of other DsbAs. Functionally, BperDsbA shows limited substrate promiscuity and selectively catalyzes the oxidative folding of a pertussis toxin-derived peptide, supporting a model of substrate specialization. Together, these findings suggest that BperDsbA has evolved unique redox and structural features to support virulence factor maturation in B. pertussis. This work expands our understanding of the mechanistic diversity of DsbA enzymes and highlights their potential as pathogen-specific targets for anti-virulence therapeutics.

Item ID:	90241
Item Type:	Article (Research - C1)
ISSN:	1469-896X
Copyright Information:	This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. © 2025 The Author(s). Protein Science published by Wiley Periodicals LLC on behalf of The Protein Society.
Funders:	National Health and Medical Research Council of Australia (NHMRC), Australian Research Council (ARC)
Projects and Grants:	NHMRC GNT1143638, NHMRC GNT1144046, ARC DP190101613, ARC DP210100673, ARC DP250102263
Date Deposited:	20 Jan 2026 01:00
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