4.7 Article

Hybrid Thermophilic/Mesophilic Enzymes Reveal a Role for Conformational Disorder in Regulation of Bacterial Enzyme I

期刊

JOURNAL OF MOLECULAR BIOLOGY
卷 432, 期 16, 页码 4481-4498

出版社

ACADEMIC PRESS LTD- ELSEVIER SCIENCE LTD
DOI: 10.1016/j.jmb.2020.05.024

关键词

enzyme regulation; bacterial PTS; metadynamics; NMR relaxation; protein dynamics

资金

  1. NIGMS [R35GM133488]
  2. Roy J. Carver Charitable Trust
  3. Intramural Research Program of the National Institutes of Health, the National Institute of Diabetes and Digestive and Kidney Disease
  4. ALS-ENABLE program - National Institutes of Health, National Institute of General Medical Sciences [DE-AC02-05CH11231, P30 GM124169-01]
  5. DOE Office of Science [DE-AC02-06CH11357]
  6. Department of Chemistry, Iowa State University
  7. NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES [ZIADK033007] Funding Source: NIH RePORTER

向作者/读者索取更多资源

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. (C) 2020 Elsevier Ltd. All rights reserved.

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