Tree-dimensional structure of the fluorescent complex of bacterial lipocalin triple mutant with chromophore – DiB3-F53L/F74L/L129M

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Abstract

The three dimensional structure of DiB3-F53L/F74L/L129M – a triple mutant of the fluorescent non-covalent complex of a genetically engineered variant of the bacterial protein lipocalin Blc with the synthetic GFP-like chromophore M739, has been studied by molecular dynamics methods. It was found that the chromophore binding site in the complex is similar to DiB1 and differs from the alternative site in DiB3. The complex is characterized by increased fluorescence brightness compared to those with other genetically engineered variants of lipocalin, which makes it one of the promising markers of biological objects in cell biology.

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About the authors

A. V. Rossokhin

Shemyakin–Ovchinnikov Institute of Bioorganic Chemistry RAS; Research Center of Neurology

Email: goryacheva@ibch.ru

Научный центр неврологии, Институт мозга

Russian Federation, ul. Miklukho-Maklaya 16/10, Moscow, 117997; per. Obuha 5, Moscow, 105064

E. A. Goryacheva

Shemyakin–Ovchinnikov Institute of Bioorganic Chemistry RAS

Author for correspondence.
Email: goryacheva@ibch.ru
Russian Federation, ul. Miklukho-Maklaya 16/10, Moscow, 117997

I. V. Artemyev

Shemyakin–Ovchinnikov Institute of Bioorganic Chemistry RAS

Email: goryacheva@ibch.ru
Russian Federation, ul. Miklukho-Maklaya 16/10, Moscow, 117997

S. F. Arhipova

Shemyakin–Ovchinnikov Institute of Bioorganic Chemistry RAS

Email: goryacheva@ibch.ru
Russian Federation, ul. Miklukho-Maklaya 16/10, Moscow, 117997

A. R. Gilvanov

Shemyakin–Ovchinnikov Institute of Bioorganic Chemistry RAS

Email: goryacheva@ibch.ru
Russian Federation, ul. Miklukho-Maklaya 16/10, Moscow, 117997

N. V. Pletneva

Shemyakin–Ovchinnikov Institute of Bioorganic Chemistry RAS

Email: goryacheva@ibch.ru
Russian Federation, ul. Miklukho-Maklaya 16/10, Moscow, 117997

V. Z. Pletnev

Shemyakin–Ovchinnikov Institute of Bioorganic Chemistry RAS

Email: goryacheva@ibch.ru
Russian Federation, ul. Miklukho-Maklaya 16/10, Moscow, 117997

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Supplementary files

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2. Fig. 1. Comparison of amino acid sequences of the parental bacterial lipocalin Blc [2] and its mutants in complexes with the synthetic GFP-like chromophore M739 [3]. Amino acid substitutions are highlighted in red. DiB3mut is a triple mutant DiB3-F53L/F74L/L129M.

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3. Fig. 2. (a) – Structural formula of the M739 chromophore; (b) – spatial structure of the lipocalin complex (shown in green) with the chromophore (shown in yellow).

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4. Fig. 3. The immediate amino acid environment of the M739 chromophore in the binding cavity of lipocalin.

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5. Fig. 4. Dependence of RMSD (Å) of the protein main chain atoms on time (iteration number 500 = 1 ns): without chromophore (a) and with the chromophore pulled along the protein cavity (b).

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6. Fig. 5. Dependence of the chromophore–protein interaction energy on time (iteration number 500 = 1 ns) during the chromophore’s movement inside the binding cavity.

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