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Synthetic Protein Switches

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Cover of 'Synthetic Protein Switches'

Table of Contents

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    Book Overview
  2. Altmetric Badge
    Chapter 1 Synthetic Protein Switches: Theoretical and Experimental Considerations
  3. Altmetric Badge
    Chapter 2 Construction of Allosteric Protein Switches by Alternate Frame Folding and Intermolecular Fragment Exchange
  4. Altmetric Badge
    Chapter 3 Construction of Protein Switches by Domain Insertion and Directed Evolution
  5. Altmetric Badge
    Chapter 4 Catalytic Amyloid Fibrils That Bind Copper to Activate Oxygen
  6. Altmetric Badge
    Chapter 5 Ancestral Protein Reconstruction and Circular Permutation for Improving the Stability and Dynamic Range of FRET Sensors
  7. Altmetric Badge
    Chapter 6 Method for Developing Optical Sensors Using a Synthetic Dye-Fluorescent Protein FRET Pair and Computational Modeling and Assessment
  8. Altmetric Badge
    Chapter 7 Rational Design and Applications of Semisynthetic Modular Biosensors: SNIFITs and LUCIDs
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    Chapter 8 Ultrasensitive Firefly Luminescent Intermediate-Based Protein-Protein Interaction Assay (FlimPIA) Based on the Functional Complementation of Mutant Firefly Luciferases
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    Chapter 9 Quantitative and Dynamic Imaging of ATM Kinase Activity
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    Chapter 10 Creation of Antigen-Dependent β-Lactamase Fusion Protein Tethered by Circularly Permuted Antibody Variable Domains
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    Chapter 11 Protein and Protease Sensing by Allosteric Derepression
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    Chapter 12 DNA-Specific Biosensors Based on Intramolecular β-Lactamase-Inhibitor Complex Formation
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    Chapter 13 Engineering and Characterizing Synthetic Protease Sensors and Switches
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    Chapter 14 Characterizing Dynamic Protein–Protein Interactions Using the Genetically Encoded Split Biosensor Assay Technique Split TEV
  16. Altmetric Badge
    Chapter 15 Development of a Synthetic Switch to Control Protein Stability in Eukaryotic Cells with Light
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    Chapter 16 Light-Regulated Protein Kinases Based on the CRY2-CIB1 System
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    Chapter 17 Yeast-Based Screening System for the Selection of Functional Light-Driven K+ Channels
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    Chapter 18 Primer-Aided Truncation for the Creation of Hybrid Proteins
  20. Altmetric Badge
    Chapter 19 Engineering Small Molecule Responsive Split Protein Kinases
  21. Altmetric Badge
    Chapter 20 Directed Evolution Methods to Rewire Signaling Networks
Attention for Chapter 5: Ancestral Protein Reconstruction and Circular Permutation for Improving the Stability and Dynamic Range of FRET Sensors
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Chapter title
Ancestral Protein Reconstruction and Circular Permutation for Improving the Stability and Dynamic Range of FRET Sensors
Chapter number 5
Book title
Synthetic Protein Switches
Published in
Methods in molecular biology, March 2017
DOI 10.1007/978-1-4939-6940-1_5
Pubmed ID
Book ISBNs
978-1-4939-6938-8, 978-1-4939-6940-1
Authors

Ben E. Clifton, Jason H. Whitfield, Inmaculada Sanchez-Romero, Michel K. Herde, Christian Henneberger, Harald Janovjak, Colin J. Jackson

Editors

Viktor Stein

Abstract

Small molecule biosensors based on Förster resonance energy transfer (FRET) enable small molecule signaling to be monitored with high spatial and temporal resolution in complex cellular environments. FRET sensors can be constructed by fusing a pair of fluorescent proteins to a suitable recognition domain, such as a member of the solute-binding protein (SBP) superfamily. However, naturally occurring SBPs may be unsuitable for incorporation into FRET sensors due to their low thermostability, which may preclude imaging under physiological conditions, or because the positions of their N- and C-termini may be suboptimal for fusion of fluorescent proteins, which may limit the dynamic range of the resulting sensors. Here, we show how these problems can be overcome using ancestral protein reconstruction and circular permutation. Ancestral protein reconstruction, used as a protein engineering strategy, leverages phylogenetic information to improve the thermostability of proteins, while circular permutation enables the termini of an SBP to be repositioned to maximize the dynamic range of the resulting FRET sensor. We also provide a protocol for cloning the engineered SBPs into FRET sensor constructs using Golden Gate assembly and discuss considerations for in situ characterization of the FRET sensors.

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Mendeley readers

Mendeley readers

The data shown below were compiled from readership statistics for 17 Mendeley readers of this research output. Click here to see the associated Mendeley record.

Geographical breakdown

Country Count As %
Unknown 17 100%

Demographic breakdown

Readers by professional status Count As %
Researcher 5 29%
Student > Bachelor 2 12%
Professor 2 12%
Other 1 6%
Student > Master 1 6%
Other 3 18%
Unknown 3 18%
Readers by discipline Count As %
Biochemistry, Genetics and Molecular Biology 7 41%
Chemistry 3 18%
Agricultural and Biological Sciences 2 12%
Neuroscience 1 6%
Unknown 4 24%
Attention Score in Context

Attention Score in Context

This research output has an Altmetric Attention Score of 1. This is our high-level measure of the quality and quantity of online attention that it has received. This Attention Score, as well as the ranking and number of research outputs shown below, was calculated when the research output was last mentioned on 05 October 2022.
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#18,951,048
of 23,482,849 outputs
Outputs from Methods in molecular biology
#8,142
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Outputs of similar age
#236,624
of 309,288 outputs
Outputs of similar age from Methods in molecular biology
#180
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