The Recombinant Inhibitor of DNA Binding Id2 Forms Multimeric Structures via the Helix-Loop-Helix Domain and the Nuclear Export Signal

Roschger, Cornelia; Schubert, Mario; Regl, Christof; Andosch, Ancuela; Marquez, Augusto; Berger, Thomas; Huber, Christian G.; Lütz-Meindl, Ursula; Cabrele, Chiara

doi:10.3390/ijms19041105

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The Recombinant Inhibitor of DNA Binding Id2 Forms Multimeric Structures via the Helix-Loop-Helix Domain and the Nuclear Export Signal

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Department of Biosciences, University of Salzburg, Billrothstrasse 11 and Hellbrunner Strasse 34, 5020 Salzburg, Austria

Christian Doppler Laboratory for Innovative Tools for Biosimilar Characterization, University of Salzburg, Hellbrunner Strasse 34, 5020 Salzburg, Austria

Department of Chemistry and Physics of Materials, University of Salzburg, Jakob-Haringer Strasse 2a, 5020 Salzburg, Austria

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Int. J. Mol. Sci. 2018, 19(4), 1105; https://doi.org/10.3390/ijms19041105

Received: 28 January 2018 / Revised: 31 March 2018 / Accepted: 3 April 2018 / Published: 7 April 2018

(This article belongs to the Section Molecular Biophysics)

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Abstract

The inhibitor of DNA binding and cell differentiation 2 (Id2) is a helix-loop-helix (HLH) protein that acts as negative dominant regulator of basic-HLH transcription factors during development and in cancer. The structural properties of Id2 have been investigated so far by using synthetic or recombinant fragments reproducing single domains (N-terminus, HLH, C-terminus): the HLH domain tends to dimerize into a four-helix bundle, whereas the flanking regions are flexible. In this work, the intact protein was expressed in E. coli, solubilized from inclusion bodies with urea, purified and dissolved in water at pH~4. Under these conditions, Id2 was obtained with both cysteine residues disulfide-bonded to β-mercaptoethanol that was present during the solubilization process. Moreover, it existed in a self-assembled state, in which the N-terminus remained highly flexible, while the HLH domain and, surprisingly, part of the C-terminus, which corresponds to the nuclear export signal (NES), both were involved in slowly tumbling, rigid structures. The protein oligomers also formed twisted fibrils that were several micrometers long and up to 80 nm thick. These results show that self-assembly decreases the backbone flexibility of those two protein regions (HLH and NES) that are important for interaction with basic-HLH transcription factors or for nucleocytoplasmic shuttling. View Full-Text

Keywords: helix-loop-helix domain; intrinsic disorder; protein oligomerization; protein self-assembly; inhibitor of DNA binding; protein fibrils

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MDPI and ACS Style

Roschger, C.; Schubert, M.; Regl, C.; Andosch, A.; Marquez, A.; Berger, T.; Huber, C.G.; Lütz-Meindl, U.; Cabrele, C. The Recombinant Inhibitor of DNA Binding Id2 Forms Multimeric Structures via the Helix-Loop-Helix Domain and the Nuclear Export Signal. Int. J. Mol. Sci. 2018, 19, 1105. https://doi.org/10.3390/ijms19041105

AMA Style

Roschger C, Schubert M, Regl C, Andosch A, Marquez A, Berger T, Huber CG, Lütz-Meindl U, Cabrele C. The Recombinant Inhibitor of DNA Binding Id2 Forms Multimeric Structures via the Helix-Loop-Helix Domain and the Nuclear Export Signal. International Journal of Molecular Sciences. 2018; 19(4):1105. https://doi.org/10.3390/ijms19041105

Chicago/Turabian Style

Roschger, Cornelia, Mario Schubert, Christof Regl, Ancuela Andosch, Augusto Marquez, Thomas Berger, Christian G. Huber, Ursula Lütz-Meindl, and Chiara Cabrele. 2018. "The Recombinant Inhibitor of DNA Binding Id2 Forms Multimeric Structures via the Helix-Loop-Helix Domain and the Nuclear Export Signal" International Journal of Molecular Sciences 19, no. 4: 1105. https://doi.org/10.3390/ijms19041105

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The Recombinant Inhibitor of DNA Binding Id2 Forms Multimeric Structures via the Helix-Loop-Helix Domain and the Nuclear Export Signal

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