Fluorescence Resonance Energy Transfer Based on Interaction of PII and PipX Proteins Provides a Robust and Specific Biosensor for 2-Oxoglutarate, a Central Metabolite and a Signaling Molecule

Chen, Hailin; Bernard, Christophe; Hubert, P.; My, Lætitia; Zhang, Cheng‐Cai

doi:10.1111/j.1742-4658.2013.12702.x

Cited by 3 publications

(2 citation statements)

References 14 publications

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“…If the probe is specific, just the selected molecule will have a significant influence on the FRET efficiency. Since most of the pre-existing biological recognition domains that are used in biosensors have high affinities and might be saturated, it should be considered to perform site-specific mutations to reduce potential saturation problems [ 33 , 34 , 35 , 36 ]. Yet not just the recognition domain plays an important role in FRET-based biosensors.…”

Section: Considerations and Limitations Of Fret Based Biosensorsmentioning

confidence: 99%

Fluorescent Proteins as Genetically Encoded FRET Biosensors in Life Sciences

Hochreiter

Garcia

Schmid

2015

Sensors

171

121

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Fluorescence- or Förster resonance energy transfer (FRET) is a measurable physical energy transfer phenomenon between appropriate chromophores, when they are in sufficient proximity, usually within 10 nm. This feature has made them incredibly useful tools for many biomedical studies on molecular interactions. Furthermore, this principle is increasingly exploited for the design of biosensors, where two chromophores are linked with a sensory domain controlling their distance and thus the degree of FRET. The versatility of these FRET-biosensors made it possible to assess a vast amount of biological variables in a fast and standardized manner, allowing not only high-throughput studies but also sub-cellular measurements of biological processes. In this review, we aim at giving an overview over the recent advances in genetically encoded, fluorescent-protein based FRET-biosensors, as these represent the largest and most vividly growing group of FRET-based sensors. For easy understanding, we are grouping them into four categories, depending on their molecular mechanism. These are based on: (a) cleavage; (b) conformational-change; (c) mechanical force and (d) changes in the micro-environment. We also address the many issues and considerations that come with the development of FRET-based biosensors, as well as the possibilities that are available to measure them.

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Section: Considerations and Limitations Of Fret Based Biosensorsmentioning

confidence: 99%

Fluorescent Proteins as Genetically Encoded FRET Biosensors in Life Sciences

Hochreiter

Garcia

Schmid

2015

Sensors

171

121

View full text Add to dashboard Cite

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“…Despite these challenges, many studies have determined the levels of 2-OG in E. coli cell extracts by applying either coupled enzymatic assays (16)(17)(18), mass spectrometry (9,(19)(20)(21), or high-pressure liquid chromatography (HPLC) methods (13). More recently, engineered fluorescent protein probes, or biosensors, have been developed to allow monitoring of 2-OG levels in cell extracts or directly inside living cells (22)(23)(24).…”

Section: Fluctuations In the 2-og Pool In Response To Nitrogen Availamentioning

confidence: 99%

The Emergence of 2-Oxoglutarate as a Master Regulator Metabolite

Huergo

Dixon

2015

Microbiol Mol Biol Rev

248

221

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SUMMARYThe metabolite 2-oxoglutarate (also known as α-ketoglutarate, 2-ketoglutaric acid, or oxoglutaric acid) lies at the intersection between the carbon and nitrogen metabolic pathways. This compound is a key intermediate of one of the most fundamental biochemical pathways in carbon metabolism, the tricarboxylic acid (TCA) cycle. In addition, 2-oxoglutarate also acts as the major carbon skeleton for nitrogen-assimilatory reactions. Experimental data support the conclusion that intracellular levels of 2-oxoglutarate fluctuate according to nitrogen and carbon availability. This review summarizes how nature has capitalized on the ability of 2-oxoglutarate to reflect cellular nutritional status through evolution of a variety of 2-oxoglutarate-sensing regulatory proteins. The number of metabolic pathways known to be regulated by 2-oxoglutarate levels has increased significantly in recent years. The signaling properties of 2-oxoglutarate are highlighted by the fact that this metabolite regulates the synthesis of the well-established master signaling molecule, cyclic AMP (cAMP), inEscherichia coli.

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Distinctive Features of PipX, a Unique Signaling Protein of Cyanobacteria

Labella

Cantos

Salinas

et al. 2020

Life

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PipX is a unique cyanobacterial protein identified by its ability to bind to PII and NtcA, two key regulators involved in the integration of signals of the nitrogen/carbon and energy status, with a tremendous impact on nitrogen assimilation and gene expression in cyanobacteria. PipX provides a mechanistic link between PII, the most widely distributed signaling protein, and NtcA, a global transcriptional regulator of cyanobacteria. PII, required for cell survival unless PipX is inactivated or down-regulated, functions by protein–protein interactions with transcriptional regulators, transporters, and enzymes. In addition, PipX appears to be involved in a wider signaling network, supported by the following observations: (i) PII–PipX complexes interact with PlmA, an as yet poorly characterized transcriptional regulator also restricted to cyanobacteria; (ii) the pipX gene is functionally connected with pipY, a gene encoding a universally conserved pyridoxal phosphate binding protein (PLPBP) involved in vitamin B6 and amino acid homeostasis, whose loss-of-function mutations cause B6-dependent epilepsy in humans, and (iii) pipX is part of a relatively robust, six-node synteny network that includes pipY and four additional genes that might also be functionally connected with pipX. In this overview, we propose that the study of the protein–protein interaction and synteny networks involving PipX would contribute to understanding the peculiarities and idiosyncrasy of signaling pathways that are conserved in cyanobacteria.

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Fluorescence Resonance Energy Transfer Based on Interaction of PII and PipX Proteins Provides a Robust and Specific Biosensor for 2-Oxoglutarate, a Central Metabolite and a Signaling Molecule

Abstract: PipX binds to PII by fluorescent resonance energy transfer (1, 2, 3) This article is protected by copyright. All rights reserved.

Cited by 3 publications

References 14 publications

Fluorescent Proteins as Genetically Encoded FRET Biosensors in Life Sciences

Fluorescent Proteins as Genetically Encoded FRET Biosensors in Life Sciences

The Emergence of 2-Oxoglutarate as a Master Regulator Metabolite

Distinctive Features of PipX, a Unique Signaling Protein of Cyanobacteria

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