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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...
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/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. |
format | Online Article Text |
id | pubmed-3468514 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2012 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
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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