Genetically encoded sensors for ions and metabolites

Okumoto, Sakiko; Deuschle, Karen; Fehr, Marcus; Hilpert, Melanie; Lager, Ida; Lalonde, Sylvie; Looger, Loren L.; Person, Jörgen; Schmidt, Anja; Frommer, Wolf B.

doi:10.1080/00380768.2004.10408560

Cited by 4 publications

(2 citation statements)

References 41 publications

(34 reference statements)

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“…In recent years, a number of sensory derivatives of fluorescent proteins have been designed. [121][122][123][124][125][126][127][128][129][130][131][132][133][134][135][136][137][138][139] These genetically encoded biosensors can complement spectroscopic data of the target molecule by probing essential cytoplasmic properties in space and time, e.g. the concentration of ions and small molecules as well as pH and reduction potential (Fig.…”

Section: Challenges and Chancesmentioning

confidence: 99%

Concepts in bio-molecular spectroscopy: vibrational case studies on metalloenzymes

Horch

Hildebrandt

Zebger

2015

Phys. Chem. Chem. Phys.

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Spectroscopic techniques play a major role in the elucidation of structure-function relationships of biological macromolecules. Here we describe an integrated approach for bio-molecular spectroscopy that takes into account the special characteristics of such compounds. The underlying fundamental concepts will be exemplarily illustrated by means of selected case studies on biocatalysts, namely hydrogenase and superoxide reductase. The treatise will be concluded with an overview of challenges and future prospects, laying emphasis on functional dynamics, in vivo studies, and computational spectroscopy.

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Section: Challenges and Chancesmentioning

confidence: 99%

Concepts in bio-molecular spectroscopy: vibrational case studies on metalloenzymes

Horch

Hildebrandt

Zebger

2015

Phys. Chem. Chem. Phys.

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“…The N and C termini move closer together upon ligand binding by 0.7 nm in case of MBP, while in the case of RBP the termini move further apart by 0.2 nm (cf. Okumoto et al 2004). The similar FRET ratio change values may thus be due to the use of flexible linkers leading to rotational averaging of the fluorophore dipole orientation ensemble.…”

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confidence: 98%

Construction and optimization of a family of genetically encoded metabolite sensors by semirational protein engineering

et al. 2005

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A family of genetically-encoded metabolite sensors has been constructed using bacterial periplasmic binding proteins (PBPs) linearly fused to protein fluorophores. The ligand-induced conformational change in a PBP allosterically regulates the relative distance and orientation of a fluorescence resonance energy transfer (FRET)-compatible protein pair. Ligand binding is transduced into a macroscopic FRET observable, providing a reagent for in vitro and in vivo ligand measurement and visualization. Sensors with a higher FRET signal change are required to expand the dynamic range and allow visualization of subtle analyte changes under high noise conditions. Various observations suggest that factors other than inter-fluorophore separation contribute to FRET transfer efficiency and the resulting ligand-dependent spectral changes. Empirical and rational protein engineering leads to enhanced allosteric linkage between ligand binding and chromophore rearrangement; modifications predicted to decrease chromophore rotational averaging enhance the signal change, emphasizing the importance of the rotational freedom parameter k 2 to FRET efficiency. Tighter allosteric linkage of the PBP and the fluorophores by linker truncation or by insertion of chromophores into the binding protein at rationally designed sites gave rise to sensors with improved signal change. High-response sensors were obtained with fluorescent proteins attached to the same binding PBP lobe, suggesting that indirect allosteric regulation during the hinge-bending motion is sufficient to give rise to a FRET response. The optimization of sensors for glucose and glutamate, ligands of great clinical interest, provides a general framework for the manipulation of ligand-dependent allosteric signal transduction mechanisms.Keywords: conformational changes; structure/function studies; new methods; molecular mechanics/ dynamics; glucose; glutamate; nanosensor; neurotransmitter; biosensor; protein engineering; FRET Supplemental material: see www.proteinscience.org Biosensors are molecular machines linking the highly evolved ligand-binding properties of a biological macromolecule to the production of a physical observable (Nakamura and Karube 2003;Looger et al. 2005). Many classes of biomolecules have been employed as ''recognition elements'' of biosensors, although proteins have emerged as the gold standard due to their superior properties, including specificity, affinity, and stability. Moreover, such sensors may be genetically encodable, thus providing

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Study of Metabolic Control in Plants by Metabolomics

Fiehn¹

Control of Primary Metabolism in Plants

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Genetically encoded sensors for ions and metabolites

Cited by 4 publications

References 41 publications

Concepts in bio-molecular spectroscopy: vibrational case studies on metalloenzymes

Concepts in bio-molecular spectroscopy: vibrational case studies on metalloenzymes

Construction and optimization of a family of genetically encoded metabolite sensors by semirational protein engineering

Study of Metabolic Control in Plants by Metabolomics

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