Caged Activators of Artificial Allosteric Protein Biosensors
Description
The ability of proteins to interconvert unrelated biochemical inputs and outputs underlays most energy and information processing in biology. A common conversion mechanism involves a conformational change of a protein receptor in response to a ligand binding or a covalent modification, leading to allosteric activity modulation of the effector domain. Designing such systems rationally is a central goal of synthetic biology and protein engineering. A two-component sensory system based on the scaffolding of modules in the presence of an analyte is one of the most generalizable biosensor architectures. An inherent problem of such systems is dependence of the response on the absolute and relative concentrations of the components. Here we use the example of two-component sensory systems based on calmodulin-operated synthetic switches to analyze and address this issue. We constructed "caged"versions of the activating domain thereby creating a thermodynamic barrier for spontaneous activation of the system. We demonstrate that the caged biosensor architectures could operate at concentrations spanning 3 orders of magnitude and are applicable to electrochemical, luminescent, and fluorescent two-component biosensors. We analyzed the activation kinetics of the caged biosensors and determined that the core allosteric switch is likely to be the rate limiting component of the system. These findings provide guidance for predictable engineering of robust sensory systems with inputs and outputs of choice.
Impact and interest:
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ID Code: | 213549 | ||||||
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Item Type: | Contribution to Journal (Journal Article) | ||||||
Refereed: | Yes | ||||||
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Additional Information: | Funding Information: This work was supported in part by the Australian Research Council Discovery Projects DP160100973 and DP150100936 as well ITTC Grant IC160100027 and NHMRC Grant APP1113262 to K.A. This work was also in part supported by HFSP Grant RGP0002/2018 to K.A. This work was also supported by contract research funding provided by Molecular Warehouse Ltd. K.A. gratefully acknowledges financial support of QUT/CSIRO Synthetic Biology Alliance. J.W. was supported by a CSIRO Synthetic Biology Future Science Fellowship from the CSIRO Synthetic Biology Future Science Platform. | ||||||
Measurements or Duration: | 9 pages | ||||||
DOI: | 10.1021/acssynbio.9b00500 | ||||||
ISSN: | 2161-5063 | ||||||
Pure ID: | 98981211 | ||||||
Divisions: | Current > Research Centres > Centre for Agriculture and the Bioeconomy Past > QUT Faculties & Divisions > Science & Engineering Faculty Current > QUT Faculties and Divisions > Faculty of Science Current > Schools > School of Biology & Environmental Science |
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Funding Information: | This work was supported in part by the Australian Research Council Discovery Projects DP160100973 and DP150100936 as well ITTC Grant IC160100027 and NHMRC Grant APP1113262 to K.A. This work was also in part supported by HFSP Grant RGP0002/2018 to K.A. This work was also supported by contract research funding provided by Molecular Warehouse Ltd. K.A. gratefully acknowledges financial support of QUT/CSIRO Synthetic Biology Alliance. J.W. was supported by a CSIRO Synthetic Biology Future Science Fellowship from the CSIRO Synthetic Biology Future Science Platform. This work was supported in part by the Australian Research Council Discovery Projects DP160100973 and DP150100936 as well ITTC Grant IC160100027 and NHMRC Grant APP1113262 to K.A. This work was also in part supported by HFSP Grant RGP0002/2018 to K.A. This work was also supported by contract research funding provided by Molecular Warehouse Ltd. K.A. gratefully acknowledges financial support of QUT/CSIRO Synthetic Biology Alliance. J.W. was supported by a CSIRO Synthetic Biology Future Science Fellowship from the CSIRO Synthetic Biology Future Science Platform. | ||||||
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Copyright Owner: | 2020 American Chemical Society | ||||||
Copyright Statement: | This work is covered by copyright. Unless the document is being made available under a Creative Commons Licence, you must assume that re-use is limited to personal use and that permission from the copyright owner must be obtained for all other uses. If the document is available under a Creative Commons License (or other specified license) then refer to the Licence for details of permitted re-use. It is a condition of access that users recognise and abide by the legal requirements associated with these rights. If you believe that this work infringes copyright please provide details by email to qut.copyright@qut.edu.au | ||||||
Deposited On: | 28 Sep 2021 02:59 | ||||||
Last Modified: | 21 Jun 2024 14:31 |
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