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Near-Infrared Markers based on Bacterial Phytochromes with Phycocyanobilin as a Chromophore

Biomarkers engineered on the basis of bacterial phytochromes with biliverdin IXα (BV) cofactor as a chromophore are increasingly used in cell biology and biomedicine, since their absorption and fluorescence spectra lie within the so-called optical “transparency window” of biological tissues. However...

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Autores principales: Stepanenko, Olesya V., Stepanenko, Olga V., Shpironok, Olesya G., Fonin, Alexander V., Kuznetsova, Irina M., Turoverov, Konstantin K.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2019
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6928796/
https://www.ncbi.nlm.nih.gov/pubmed/31810174
http://dx.doi.org/10.3390/ijms20236067
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author Stepanenko, Olesya V.
Stepanenko, Olga V.
Shpironok, Olesya G.
Fonin, Alexander V.
Kuznetsova, Irina M.
Turoverov, Konstantin K.
author_facet Stepanenko, Olesya V.
Stepanenko, Olga V.
Shpironok, Olesya G.
Fonin, Alexander V.
Kuznetsova, Irina M.
Turoverov, Konstantin K.
author_sort Stepanenko, Olesya V.
collection PubMed
description Biomarkers engineered on the basis of bacterial phytochromes with biliverdin IXα (BV) cofactor as a chromophore are increasingly used in cell biology and biomedicine, since their absorption and fluorescence spectra lie within the so-called optical “transparency window” of biological tissues. However, the quantum yield of BV fluorescence in these biomarkers does not exceed 0.145. The task of generating biomarkers with a higher fluorescence quantum yield remains relevant. To address the problem, we proposed the use of phycocyanobilin (PCB) as a chromophore of biomarkers derived from bacterial phytochromes. In this work, we characterized the complexes of iRFP713 evolved from RpBphP2 and its mutant variants with different location of cysteine residues capable of covalent tetrapyrrole attachment with the PCB cofactor. All analyzed proteins assembled with PCB were shown to have a higher fluorescence quantum yield than the proteins assembled with BV. The iRFP713/V256C and iRFP713/C15S/V256C assembled with PCB have a particularly high quantum yield of 0.5 and 0.45, which exceeds the quantum yield of all currently available near-infrared biomarkers. Moreover, PCB has 4 times greater affinity for iRFP713/V256C and iRFP713/C15S/V256C proteins compared to BV. These data establish iRFP713/V256C and iRFP713/C15S/V256C assembled with the PCB chromophore as promising biomarkers for application in vivo. The analysis of the spectral properties of the tested biomarkers allowed for suggesting that the high-fluorescence quantum yield of the PCB chromophore can be attributed to the lower mobility of the D-ring of PCB compared to BV.
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spelling pubmed-69287962019-12-26 Near-Infrared Markers based on Bacterial Phytochromes with Phycocyanobilin as a Chromophore Stepanenko, Olesya V. Stepanenko, Olga V. Shpironok, Olesya G. Fonin, Alexander V. Kuznetsova, Irina M. Turoverov, Konstantin K. Int J Mol Sci Article Biomarkers engineered on the basis of bacterial phytochromes with biliverdin IXα (BV) cofactor as a chromophore are increasingly used in cell biology and biomedicine, since their absorption and fluorescence spectra lie within the so-called optical “transparency window” of biological tissues. However, the quantum yield of BV fluorescence in these biomarkers does not exceed 0.145. The task of generating biomarkers with a higher fluorescence quantum yield remains relevant. To address the problem, we proposed the use of phycocyanobilin (PCB) as a chromophore of biomarkers derived from bacterial phytochromes. In this work, we characterized the complexes of iRFP713 evolved from RpBphP2 and its mutant variants with different location of cysteine residues capable of covalent tetrapyrrole attachment with the PCB cofactor. All analyzed proteins assembled with PCB were shown to have a higher fluorescence quantum yield than the proteins assembled with BV. The iRFP713/V256C and iRFP713/C15S/V256C assembled with PCB have a particularly high quantum yield of 0.5 and 0.45, which exceeds the quantum yield of all currently available near-infrared biomarkers. Moreover, PCB has 4 times greater affinity for iRFP713/V256C and iRFP713/C15S/V256C proteins compared to BV. These data establish iRFP713/V256C and iRFP713/C15S/V256C assembled with the PCB chromophore as promising biomarkers for application in vivo. The analysis of the spectral properties of the tested biomarkers allowed for suggesting that the high-fluorescence quantum yield of the PCB chromophore can be attributed to the lower mobility of the D-ring of PCB compared to BV. MDPI 2019-12-02 /pmc/articles/PMC6928796/ /pubmed/31810174 http://dx.doi.org/10.3390/ijms20236067 Text en © 2019 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Stepanenko, Olesya V.
Stepanenko, Olga V.
Shpironok, Olesya G.
Fonin, Alexander V.
Kuznetsova, Irina M.
Turoverov, Konstantin K.
Near-Infrared Markers based on Bacterial Phytochromes with Phycocyanobilin as a Chromophore
title Near-Infrared Markers based on Bacterial Phytochromes with Phycocyanobilin as a Chromophore
title_full Near-Infrared Markers based on Bacterial Phytochromes with Phycocyanobilin as a Chromophore
title_fullStr Near-Infrared Markers based on Bacterial Phytochromes with Phycocyanobilin as a Chromophore
title_full_unstemmed Near-Infrared Markers based on Bacterial Phytochromes with Phycocyanobilin as a Chromophore
title_short Near-Infrared Markers based on Bacterial Phytochromes with Phycocyanobilin as a Chromophore
title_sort near-infrared markers based on bacterial phytochromes with phycocyanobilin as a chromophore
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6928796/
https://www.ncbi.nlm.nih.gov/pubmed/31810174
http://dx.doi.org/10.3390/ijms20236067
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