Thermodynamic Stability of the Transcription Regulator PaaR2 from Escherichia coli O157:H7.

TitleThermodynamic Stability of the Transcription Regulator PaaR2 from Escherichia coli O157:H7.
Publication TypeJournal Article
Year of Publication2019
AuthorsDe Bruyn, P., S. Hadži, A. Vandervelde, A. Konijnenberg, M. Prolič-Kalinšek, Y. G. - J. Sterckx, F. Sobott, J. Lah, L. Van Melderen, and R. Loris
JournalBiophys J
Volume116
Issue8
Pagination1420-1431
Date Published2019 Apr 23
ISSN1542-0086
Abstract

PaaR2 is a putative transcription regulator encoded by a three-component parDE-like toxin-antitoxin module from Escherichia coli O157:H7. Although this module's toxin, antitoxin, and toxin-antitoxin complex have been more thoroughly investigated, little remains known about its transcription regulator PaaR2. Using a wide range of biophysical techniques (circular dichroism spectroscopy, size-exclusion chromatography-multiangle laser light scattering, dynamic light scattering, small-angle x-ray scattering, and native mass spectrometry), we demonstrate that PaaR2 mainly consists of α-helices and displays a concentration-dependent octameric build-up in solution and that this octamer contains a global shape that is significantly nonspherical. Thermal unfolding of PaaR2 is reversible and displays several transitions, suggesting a complex unfolding mechanism. The unfolding data obtained from spectroscopic and calorimetric methods were combined into a unifying thermodynamic model, which suggests a five-state unfolding trajectory. Furthermore, the model allows the calculation of a stability phase diagram, which shows that, under physiological conditions, PaaR2 mainly exists as a dimer that can swiftly oligomerize into an octamer depending on local protein concentrations. These findings, based on a thorough biophysical and thermodynamic analysis of PaaR2, may provide important insights into biological function such as DNA binding and transcriptional regulation.

DOI10.1016/j.bpj.2019.03.015
Alternate JournalBiophys. J.
PubMed ID30979547
PubMed Central IDPMC6486494
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