Dynamics of a family of cyan fluorescent proteins probed by incoherent neutron scattering

Author:

Golub Maksym12,Guillon Virginia2,Gotthard Guillaume3ORCID,Zeller Dominik14,Martinez Nicolas12,Seydel Tilo1ORCID,Koza Michael M.1ORCID,Lafaye Céline2ORCID,Clavel Damien2ORCID,von Stetten David3ORCID,Royant Antoine23ORCID,Peters Judith14ORCID

Affiliation:

1. Institut Laue Langevin, 71 avenue des Martyrs, 38042 Grenoble Cedex 9, France

2. Univ. Grenoble Alpes, CNRS, CEA, IBS (Institut de Biologie Structurale), 38000 Grenoble, France

3. European Synchrotron Radiation Facility, 38043 Grenoble, France

4. Laboratoire Interdisciplinaire de Physique, Univ. Grenoble Alpes, CNRS, 38000 Grenoble, France

Abstract

Cyan fluorescent proteins (CFPs) are variants of green fluorescent proteins in which the central tyrosine of the chromophore has been replaced by a tryptophan. The increased bulk of the chromophore within a compact protein and the change in the positioning of atoms capable of hydrogen bonding have made it difficult to optimize their fluorescence properties, which took approximately 15 years between the availability of the first useable CFP, enhanced cyan fluorescent protein (ECFP), and that of a variant with almost perfect fluorescence efficiency, mTurquoise2. To understand the molecular bases of the progressive improvement in between these two CFPs, we have studied by incoherent neutron scattering the dynamics of five different variants exhibiting progressively increased fluorescence efficiency along the evolution pathway. Our results correlate well with the analysis of the previously determined X-ray crystallographic structures, which show an increase in flexibility between ECFP and the second variant, Cerulean, which is then hindered in the three later variants, SCFP3A (Super Cyan Fluorescent Protein 3A), mTurquoise and mTurquoise2. This confirms that increasing the rigidity of the direct environment of the fluorescent chromophore is not the sole parameter leading to brighter fluorescent proteins and that increased flexibility in some cases may be helpful.

Funder

Agence Nationale de Recherche

Publisher

The Royal Society

Subject

Biomedical Engineering,Biochemistry,Biomaterials,Bioengineering,Biophysics,Biotechnology

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