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Download fileCaged Activators of Artificial Allosteric Protein Biosensors
journal contribution
posted on 14.05.2020, 13:09 by Selvakumar Edwardraja, Zhong Guo, Jason Whitfield, Ignacio Retamal Lantadilla, Wayne A. Johnston, Patricia Walden, Claudia E. Vickers, Kirill AlexandrovThe
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.
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Keywords
effector domain3 ordersallosteric activity modulationCaged Activatorsbiosensor architecturesprotein receptoroutputs underlaysactivation kineticscore allosteric switchprotein engineeringgeneralizable biosensor architecturesligand bindingcovalent modificationconversion mechanisminformation processingArtificial Allosteric Protein Biosensors