Amino acid residue at the 165th position tunes EYFP chromophore maturation. A structure-based design.

Autor:	Pletneva NV; Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Moscow 117997, Russia., Maksimov EG; Faculty of Biology, Lomonosov Moscow State University, 119992 Moscow, Russia., Protasova EA; Faculty of Biology, Lomonosov Moscow State University, 119992 Moscow, Russia., Mamontova AV; Center of Life Sciences, Skolkovo Institute of Science and Technology, Moscow 121205, Russia., Simonyan TR; Center of Life Sciences, Skolkovo Institute of Science and Technology, Moscow 121205, Russia., Ziganshin RH; Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Moscow 117997, Russia., Lukyanov KA; Center of Life Sciences, Skolkovo Institute of Science and Technology, Moscow 121205, Russia., Muslinkina L; Structural Biology Section, Research Technologies Branch, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA., Pletnev S; Vaccine Research Center, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA., Bogdanov AM; Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Moscow 117997, Russia., Pletnev VZ; Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Moscow 117997, Russia.
Jazyk:	angličtina
Zdroj:	Computational and structural biotechnology journal [Comput Struct Biotechnol J] 2021 May 11; Vol. 19, pp. 2950-2959. Date of Electronic Publication: 2021 May 11 (Print Publication: 2021).
DOI:	10.1016/j.csbj.2021.05.017
Abstrakt:	For the whole GFP family, a few cases, when a single mutation in the chromophore environment strongly inhibits maturation, were described. Here we study EYFP-F165G - a variant of the enhanced yellow fluorescent protein - obtained by a single F165G replacement, and demonstrated multiple fluorescent states represented by the minor emission peaks in blue and yellow ranges (~470 and ~530 nm), and the major peak at ~330 nm. The latter has been assigned to tryptophan fluorescence, quenched due to excitation energy transfer to the mature chromophore in the parental EYFP protein. EYFP-F165G crystal structure revealed two general independent routes of post-translational chemistry, resulting in two main states of the polypeptide chain with the intact chromophore forming triad (~85%) and mature chromophore (~15%). Our experiments thus highlighted important stereochemical role of the 165th position strongly affecting spectral characteristics of the protein. On the basis of the determined EYFP-F165G three-dimensional structure, new variants with ~ 2-fold improved brightness were engineered. Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. (© 2021 The Authors.)
Databáze:	MEDLINE
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