ArticleProtein science : a publication of the Protein Society2015
Construction of a robust and sensitive arginine biosensor through ancestral protein reconstruction.
Article in Protein science : a publication of the Protein Society, 2015. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 24 papers.
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24 citing papers in PubMed, 72 citations in OpenAlex.
- Highly efficient production ofSynthetic and systems biotechnology · 2027Article
- Thermostable ancestors enabled evolutionary diversification of promiscuous chemical defence enzymes.The EMBO journal · 2026Article
- Engineering functional thermostable proteins using ancestral sequence reconstruction.The Journal of biological chemistry · 2022Review
- Monitoring extracellular ion and metabolite dynamics with recombinant nanobody-fused biosensors.iScience · 2022Article
- Computational design and experimental characterisation of a stable human heparanase variant.RSC chemical biology · 2022Article
- The role of C-terminal helix in the conformational transition of an arginine binding protein.Journal of structural biology: X · 2022Article
- Immobilization of Recombinant Fluorescent Biosensors Permits Imaging of Extracellular Ion Signals.ACS sensors · 2021Article
- FRET-Based Genetically Encoded Nanosensor for Real-Time Monitoring of the Flux of α-Tocopherol in Living Cells.ACS omega · 2021Article
- Engineering of Ancestors as a Tool to Elucidate Structure, Mechanism, and Specificity of Extant Terpene Cyclase.Journal of the American Chemical Society · 2021Article
- LTP Induction Boosts Glutamate Spillover by Driving Withdrawal of Perisynaptic Astroglia.Neuron · 2020Article
- Altered conformational sampling along an evolutionary trajectory changes the catalytic activity of an enzyme.Nature communications · 2020Article
- Development of a Protein Scaffold for Arginine Sensing Generated through the Dissection of the Arginine-Binding Protein fromInternational journal of molecular sciences · 2020Article
- The Role of Amino Acids in Neurotransmission and Fluorescent Tools for Their Detection.International journal of molecular sciences · 2020Review
- Ancestral Resurrection and Directed Evolution of Fungal Mesozoic Laccases.Applied and environmental microbiology · 2020Article
- Metabolic Flux Analysis of Catechin Biosynthesis Pathways Using Nanosensor.Antioxidants (Basel, Switzerland) · 2020Article
- Real-Time Optical Detection of Isoleucine in Living Cells through a Genetically-Encoded Nanosensor.Sensors (Basel, Switzerland) · 2019Article
- Highly thermostable carboxylic acid reductases generated by ancestral sequence reconstruction.Communications biology · 2019Article
- Monitoring hippocampal glycine with the computationally designed optical sensor GlyFS.Nature chemical biology · 2018Article
- Redesign of LAOBP to bind novel l-amino acid ligands.Protein science : a publication of the Protein Society · 2018Article
- An improvedThe Journal of biological chemistry · 2018Article
Corrections and comments
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Authors and funding
8 authors at 4 institutions in 4 countries.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Biosensors for signaling molecules allow the study of physiological processes by bringing together the fields of protein engineering, fluorescence imaging, and cell biology. Construction of genetically encoded biosensors generally relies on the availability of a binding "core" that is both specific and stable, which can then be combined with fluorescent molecules to create a sensor. However, binding proteins with the desired properties are often not available in nature and substantial improvement to sensors can be required, particularly with regard to their durability. Ancestral protein reconstruction is a powerful protein-engineering tool able to generate highly stable and functional proteins. In this work, we sought to establish the utility of ancestral protein reconstruction to biosensor development, beginning with the construction of an l-arginine biosensor. l-arginine, as the immediate precursor to nitric oxide, is an important molecule in many physiological contexts including brain function. Using a combination of ancestral reconstruction and circular permutation, we constructed a Förster resonance energy transfer (FRET) biosensor for l-arginine (cpFLIPR). cpFLIPR displays high sensitivity and specificity, with a Kd of ∼14 µM and a maximal dynamic range of 35%. Importantly, cpFLIPR was highly robust, enabling accurate l-arginine measurement at physiological temperatures. We established that cpFLIPR is compatible with two-photon excitation fluorescence microscopy and report l-arginine concentrations in brain tissue.
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Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.