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A Green Fluorescent Protein Containing a QFG Tri-Peptide Chromophore: Optical Properties and X-Ray Crystal Structure

Rtms5 is an deep blue weakly fluorescent GFP-like protein ([Image: see text], 592 nm; [Image: see text], 630nm; Φ(F), 0.004) that contains a (66)Gln-Tyr-Gly chromophore tripeptide sequence. We investigated the optical properties and structure of two variants, Rtms5(Y67F) and Rtms5(Y67F/H146S) in whi...

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Autores principales: Battad, Jion M., Traore, Daouda A. K., Byres, Emma, Rossjohn, Jamie, Devenish, Rodney J., Olsen, Seth, Wilce, Matthew C. J., Prescott, Mark
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2012
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3468514/
https://www.ncbi.nlm.nih.gov/pubmed/23071789
http://dx.doi.org/10.1371/journal.pone.0047331
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author Battad, Jion M.
Traore, Daouda A. K.
Byres, Emma
Rossjohn, Jamie
Devenish, Rodney J.
Olsen, Seth
Wilce, Matthew C. J.
Prescott, Mark
author_facet Battad, Jion M.
Traore, Daouda A. K.
Byres, Emma
Rossjohn, Jamie
Devenish, Rodney J.
Olsen, Seth
Wilce, Matthew C. J.
Prescott, Mark
author_sort Battad, Jion M.
collection PubMed
description Rtms5 is an deep blue weakly fluorescent GFP-like protein ([Image: see text], 592 nm; [Image: see text], 630nm; Φ(F), 0.004) that contains a (66)Gln-Tyr-Gly chromophore tripeptide sequence. We investigated the optical properties and structure of two variants, Rtms5(Y67F) and Rtms5(Y67F/H146S) in which the tyrosine at position 67 was substituted by a phenylalanine. Compared to the parent proteins the optical spectra for these new variants were significantly blue-shifted. Rtms5(Y67F) spectra were characterised by two absorbing species ([Image: see text], 440 nm and 513 nm) and green fluorescence emission ([Image: see text], 440 nm; [Image: see text], 508 nm; Φ(F), 0.11), whilst Rtms5(Y67F/H146S) spectra were characterised by a single absorbing species ([Image: see text], 440 nm) and a relatively high fluorescence quantum yield (Φ(F,) 0.75; [Image: see text], 440 nm; [Image: see text], 508 nm). The fluorescence emissions of each variant were remarkably stable over a wide range of pH (3–11). These are the first GFP-like proteins with green emissions (500–520 nm) that do not have a tyrosine at position 67. The X-ray crystal structure of each protein was determined to 2.2 Å resolution and showed that the benzylidine ring of the chromophore, similar to the 4-hydroxybenzylidine ring of the Rtms5 parent, is non-coplanar and in the trans conformation. The results of chemical quantum calculations together with the structural data suggested that the 513 nm absorbing species in Rtms5(Y67F) results from an unusual form of the chromophore protonated at the acylimine oxygen. These are the first X-ray crystal structures for fluorescent proteins with a functional chromophore containing a phenylalanine at position 67.
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spelling pubmed-34685142012-10-15 A Green Fluorescent Protein Containing a QFG Tri-Peptide Chromophore: Optical Properties and X-Ray Crystal Structure Battad, Jion M. Traore, Daouda A. K. Byres, Emma Rossjohn, Jamie Devenish, Rodney J. Olsen, Seth Wilce, Matthew C. J. Prescott, Mark PLoS One Research Article Rtms5 is an deep blue weakly fluorescent GFP-like protein ([Image: see text], 592 nm; [Image: see text], 630nm; Φ(F), 0.004) that contains a (66)Gln-Tyr-Gly chromophore tripeptide sequence. We investigated the optical properties and structure of two variants, Rtms5(Y67F) and Rtms5(Y67F/H146S) in which the tyrosine at position 67 was substituted by a phenylalanine. Compared to the parent proteins the optical spectra for these new variants were significantly blue-shifted. Rtms5(Y67F) spectra were characterised by two absorbing species ([Image: see text], 440 nm and 513 nm) and green fluorescence emission ([Image: see text], 440 nm; [Image: see text], 508 nm; Φ(F), 0.11), whilst Rtms5(Y67F/H146S) spectra were characterised by a single absorbing species ([Image: see text], 440 nm) and a relatively high fluorescence quantum yield (Φ(F,) 0.75; [Image: see text], 440 nm; [Image: see text], 508 nm). The fluorescence emissions of each variant were remarkably stable over a wide range of pH (3–11). These are the first GFP-like proteins with green emissions (500–520 nm) that do not have a tyrosine at position 67. The X-ray crystal structure of each protein was determined to 2.2 Å resolution and showed that the benzylidine ring of the chromophore, similar to the 4-hydroxybenzylidine ring of the Rtms5 parent, is non-coplanar and in the trans conformation. The results of chemical quantum calculations together with the structural data suggested that the 513 nm absorbing species in Rtms5(Y67F) results from an unusual form of the chromophore protonated at the acylimine oxygen. These are the first X-ray crystal structures for fluorescent proteins with a functional chromophore containing a phenylalanine at position 67. Public Library of Science 2012-10-10 /pmc/articles/PMC3468514/ /pubmed/23071789 http://dx.doi.org/10.1371/journal.pone.0047331 Text en © 2012 Battad 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
Battad, Jion M.
Traore, Daouda A. K.
Byres, Emma
Rossjohn, Jamie
Devenish, Rodney J.
Olsen, Seth
Wilce, Matthew C. J.
Prescott, Mark
A Green Fluorescent Protein Containing a QFG Tri-Peptide Chromophore: Optical Properties and X-Ray Crystal Structure
title A Green Fluorescent Protein Containing a QFG Tri-Peptide Chromophore: Optical Properties and X-Ray Crystal Structure
title_full A Green Fluorescent Protein Containing a QFG Tri-Peptide Chromophore: Optical Properties and X-Ray Crystal Structure
title_fullStr A Green Fluorescent Protein Containing a QFG Tri-Peptide Chromophore: Optical Properties and X-Ray Crystal Structure
title_full_unstemmed A Green Fluorescent Protein Containing a QFG Tri-Peptide Chromophore: Optical Properties and X-Ray Crystal Structure
title_short A Green Fluorescent Protein Containing a QFG Tri-Peptide Chromophore: Optical Properties and X-Ray Crystal Structure
title_sort green fluorescent protein containing a qfg tri-peptide chromophore: optical properties and x-ray crystal structure
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3468514/
https://www.ncbi.nlm.nih.gov/pubmed/23071789
http://dx.doi.org/10.1371/journal.pone.0047331
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