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Mining the Sinorhizobium meliloti Transportome to Develop FRET Biosensors for Sugars, Dicarboxylates and Cyclic Polyols
BACKGROUND: Förster resonance energy transfer (FRET) biosensors are powerful tools to detect biologically important ligands in real time. Currently FRET bisosensors are available for twenty-two compounds distributed in eight classes of chemicals (two pentoses, two hexoses, two disaccharides, four am...
Autores principales: | , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Public Library of Science
2012
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3454389/ https://www.ncbi.nlm.nih.gov/pubmed/23028462 http://dx.doi.org/10.1371/journal.pone.0043578 |
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author | Bourdès, Alexandre Rudder, Steven East, Alison K. Poole, Philip S. |
author_facet | Bourdès, Alexandre Rudder, Steven East, Alison K. Poole, Philip S. |
author_sort | Bourdès, Alexandre |
collection | PubMed |
description | BACKGROUND: Förster resonance energy transfer (FRET) biosensors are powerful tools to detect biologically important ligands in real time. Currently FRET bisosensors are available for twenty-two compounds distributed in eight classes of chemicals (two pentoses, two hexoses, two disaccharides, four amino acids, one nucleobase, two nucleotides, six ions and three phytoestrogens). To expand the number of available FRET biosensors we used the induction profile of the Sinorhizobium meliloti transportome to systematically screen for new FRET biosensors. METHODOLOGY/PRINCIPAL FINDINGS: Two new vectors were developed for cloning genes for solute-binding proteins (SBPs) between those encoding FRET partner fluorescent proteins. In addition to a vector with the widely used cyan and yellow fluorescent protein FRET partners, we developed a vector using orange (mOrange2) and red fluorescent protein (mKate2) FRET partners. From the sixty-nine SBPs tested, seven gave a detectable FRET signal change on binding substrate, resulting in biosensors for D-quinic acid, myo-inositol, L-rhamnose, L-fucose, β-diglucosides (cellobiose and gentiobiose), D-galactose and C4-dicarboxylates (malate, succinate, oxaloacetate and fumarate). To our knowledge, we describe the first two FRET biosensor constructs based on SBPs from Tripartite ATP-independent periplasmic (TRAP) transport systems. CONCLUSIONS/SIGNIFICANCE: FRET based on orange (mOrange2) and red fluorescent protein (mKate2) partners allows the use of longer wavelength light, enabling deeper penetration of samples at lower energy and increased resolution with reduced back-ground auto-fluorescence. The FRET biosensors described in this paper for four new classes of compounds; (i) cyclic polyols, (ii) L-deoxy sugars, (iii) β-linked disaccharides and (iv) C4-dicarboxylates could be developed to study metabolism in vivo. |
format | Online Article Text |
id | pubmed-3454389 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2012 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-34543892012-10-01 Mining the Sinorhizobium meliloti Transportome to Develop FRET Biosensors for Sugars, Dicarboxylates and Cyclic Polyols Bourdès, Alexandre Rudder, Steven East, Alison K. Poole, Philip S. PLoS One Research Article BACKGROUND: Förster resonance energy transfer (FRET) biosensors are powerful tools to detect biologically important ligands in real time. Currently FRET bisosensors are available for twenty-two compounds distributed in eight classes of chemicals (two pentoses, two hexoses, two disaccharides, four amino acids, one nucleobase, two nucleotides, six ions and three phytoestrogens). To expand the number of available FRET biosensors we used the induction profile of the Sinorhizobium meliloti transportome to systematically screen for new FRET biosensors. METHODOLOGY/PRINCIPAL FINDINGS: Two new vectors were developed for cloning genes for solute-binding proteins (SBPs) between those encoding FRET partner fluorescent proteins. In addition to a vector with the widely used cyan and yellow fluorescent protein FRET partners, we developed a vector using orange (mOrange2) and red fluorescent protein (mKate2) FRET partners. From the sixty-nine SBPs tested, seven gave a detectable FRET signal change on binding substrate, resulting in biosensors for D-quinic acid, myo-inositol, L-rhamnose, L-fucose, β-diglucosides (cellobiose and gentiobiose), D-galactose and C4-dicarboxylates (malate, succinate, oxaloacetate and fumarate). To our knowledge, we describe the first two FRET biosensor constructs based on SBPs from Tripartite ATP-independent periplasmic (TRAP) transport systems. CONCLUSIONS/SIGNIFICANCE: FRET based on orange (mOrange2) and red fluorescent protein (mKate2) partners allows the use of longer wavelength light, enabling deeper penetration of samples at lower energy and increased resolution with reduced back-ground auto-fluorescence. The FRET biosensors described in this paper for four new classes of compounds; (i) cyclic polyols, (ii) L-deoxy sugars, (iii) β-linked disaccharides and (iv) C4-dicarboxylates could be developed to study metabolism in vivo. Public Library of Science 2012-09-24 /pmc/articles/PMC3454389/ /pubmed/23028462 http://dx.doi.org/10.1371/journal.pone.0043578 Text en © 2012 Bourdès et al http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited. |
spellingShingle | Research Article Bourdès, Alexandre Rudder, Steven East, Alison K. Poole, Philip S. Mining the Sinorhizobium meliloti Transportome to Develop FRET Biosensors for Sugars, Dicarboxylates and Cyclic Polyols |
title | Mining the Sinorhizobium meliloti Transportome to Develop FRET Biosensors for Sugars, Dicarboxylates and Cyclic Polyols |
title_full | Mining the Sinorhizobium meliloti Transportome to Develop FRET Biosensors for Sugars, Dicarboxylates and Cyclic Polyols |
title_fullStr | Mining the Sinorhizobium meliloti Transportome to Develop FRET Biosensors for Sugars, Dicarboxylates and Cyclic Polyols |
title_full_unstemmed | Mining the Sinorhizobium meliloti Transportome to Develop FRET Biosensors for Sugars, Dicarboxylates and Cyclic Polyols |
title_short | Mining the Sinorhizobium meliloti Transportome to Develop FRET Biosensors for Sugars, Dicarboxylates and Cyclic Polyols |
title_sort | mining the sinorhizobium meliloti transportome to develop fret biosensors for sugars, dicarboxylates and cyclic polyols |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3454389/ https://www.ncbi.nlm.nih.gov/pubmed/23028462 http://dx.doi.org/10.1371/journal.pone.0043578 |
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