Role of the disulfide bond in stabilizing and folding of the fimbrial protein DraE from uropathogenic Escherichia coli

Justyna Pilipczuk , Beata Zalewska-Piątek , Piotr Bruździak , Jacek Czub , Miłosz Wieczór , Marcin Olszewski , Marta Wanarska , Bogdan Nowicki , Danuta Augustin-Nowacka , Rafał Piątek


Dr fimbriae are homopolymeric adhesive organelles of uropathogenic Escherichia coli composed of DraE subunits, responsible for the attachment to host cells. These structures are characterized by enormously high stability resulting from the structural properties of an Ig-like fold of DraE. One feature of DraE and other fimbrial subunits that makes them peculiar among Ig-like domain-containing proteins is a conserved disulfide bond that joins their A and B strands. Here, we investigated how this disulfide bond affects the stability and folding/unfolding pathway of DraE. We found that the disulfide bond stabilizes self-complemented DraE (DraE-sc) by ca. 50 kJ mol-1 in an exclusively thermodynamic manner, i.e. by lowering the free energy of the native state and with almost no effect on the free energy of the transition state. This finding was confirmed by experimentally determined folding and unfolding rate constants of DraE-sc and a disulfide bond-lacking DraE-sc variant. While the folding of both proteins exhibited similar kinetics, the unfolding rate constant changed upon deletion of the disulfide bond by 10 orders of magnitude, from ca. 10-17 s-1 to 10-7 s-1. Molecular simulations revealed that unfolding of the disulfide bond-lacking variant is initiated by strands A or G and that disulfide bond-mediated joining of strand A to the core strand B cooperatively stabilizes the whole protein. We also show that the disulfide bond in DraE is recognized by the DraB chaperone, indicating a mechanism that precludes the incorporation of less stable, non oxidized DraE forms into the fimbriae.
Author Justyna Pilipczuk
Justyna Pilipczuk,,
, Beata Zalewska-Piątek
Beata Zalewska-Piątek ,,
, Piotr Bruździak
Piotr Bruździak,,
, Jacek Czub
Jacek Czub,,
, Miłosz Wieczór
Miłosz Wieczór,,
, Marcin Olszewski
Marcin Olszewski,,
, Marta Wanarska
Marta Wanarska,,
, Bogdan Nowicki
Bogdan Nowicki,,
, Danuta Augustin-Nowacka (FCh / PChW)
Danuta Augustin-Nowacka,,
- Physics-Chemistry Workshops
, Rafał Piątek
Rafał Piątek,,
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Journal seriesJournal of Biological Chemistry, ISSN 0021-9258, (A 35 pkt)
Issue year2017
Publication size in sheets0.65
ASJC Classification1303 Biochemistry; 1307 Cell Biology; 1312 Molecular Biology
Languageen angielski
LicenseJournal (articles only); published final; Uznanie Autorstwa (CC-BY); with publication
Score (nominal)35
Score sourcejournalList
ScoreMinisterial score = 35.0, 31-01-2020, ArticleFromJournal
Publication indicators WoS Citations = 2; Scopus SNIP (Source Normalised Impact per Paper): 2017 = 1.096; WoS Impact Factor: 2017 = 4.01 (2) - 2017=4.253 (5)
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