Campus Units

Biochemistry, Biophysics and Molecular Biology, Roy J. Carver Department of, Chemistry, Office of Biotechnology

Document Type

Article

Publication Version

Accepted Manuscript

Publication Date

6-3-2020

Journal or Book Title

Journal of Molecular Biology

DOI

10.1016/j.jmb.2020.05.024

Abstract

Conformational disorder is emerging as an important feature of biopolymers, regulating a vast array of cellular functions, including signaling, phase separation, and enzyme catalysis. Here we combine NMR, crystallography, computer simulations, protein engineering, and functional assays to investigate the role played by conformational heterogeneity in determining the activity of the C-terminal domain of bacterial Enzyme I (EIC). In particular, we design chimeric proteins by hybridizing EIC from thermophilic and mesophilic organisms, and we characterize the resulting constructs for structure, dynamics, and biological function. We show that EIC exists as a mixture of active and inactive conformations and that functional regulation is achieved by tuning the thermodynamic balance between active and inactive states. Interestingly, we also present a hybrid thermophilic/mesophilic enzyme that is thermostable and more active than the wild-type thermophilic enzyme, suggesting that hybridizing thermophilic and mesophilic proteins is a valid strategy to engineer thermostable enzymes with significant low-temperature activity.

Comments

This is a manuscript of an article published as Dotas, Rochelle R., Trang T. Nguyen, Charles E. Stewart Jr, Rodolfo Ghirlando, Davit A. Potoyan, and Vincenzo Venditti. "Hybrid thermophilic/mesophilic enzymes reveal a role for conformational disorder in regulation of bacterial Enzyme I." Journal of Molecular Biology (2020). DOI: 10.1016/j.jmb.2020.05.024.

Rights

Works produced by employees of the U.S. Government as part of their official duties are not copyrighted within the U.S. The content of this document is not copyrighted.

Language

en

File Format

application/pdf

Published Version

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