Мельникова Дарья Николаевна

Избранные публикации

  1. Shenkarev Z.O., Melnikova D.N., Finkina E.I., Sukhanov S.V., Boldyrev I.A., Gizatullina A.K., Mineev K.S., Arseniev A.S., Ovchinnikova T.V. (2017). Ligand Binding Properties of the Lentil Lipid Transfer Protein: Molecular Insight into the Possible Mechanism of Lipid Uptake. Biochemistry 56 (12), 1785–1796 [+]

    The lentil lipid transfer protein, designated as Lc-LTP2, was isolated from Lens culinaris seeds. The protein belongs to the LTP1 subfamily and consists of 93 amino acid residues. Its spatial structure includes four α-helices (H1-H4) and a long C-terminal tail. Here, we report the ligand binding properties of Lc-LTP2. The fluorescent 2-p-toluidinonaphthalene-6-sulfonate binding assay revealed that the affinity of Lc-LTP2 for saturated and unsaturated fatty acids was enhanced with a decrease in acyl-chain length. Measurements of boundary potential in planar lipid bilayers and calcein dye leakage in vesicular systems revealed preferential interaction of Lc-LTP2 with the negatively charged membranes. Lc-LTP2 more efficiently transferred anionic dimyristoylphosphatidylglycerol (DMPG) than zwitterionic dimyristoylphosphatidylcholine. Nuclear magnetic resonance experiments confirmed the higher affinity of Lc-LTP2 for anionic lipids and those with smaller volumes of hydrophobic chains. The acyl chains of the bound lysopalmitoylphosphatidylglycerol (LPPG), DMPG, or dihexanoylphosphatidylcholine molecules occupied the internal hydrophobic cavity, while their headgroups protruded into the aqueous environment between helices H1 and H3. The spatial structure and backbone dynamics of the Lc-LTP2-LPPG complex were determined. The internal cavity was expanded from ∼600 to ∼1000 Å(3) upon the ligand binding. Another entrance into the internal cavity, restricted by the H2-H3 interhelical loop and C-terminal tail, appeared to be responsible for the attachment of Lc-LTP2 to the membrane or micelle surface and probably played an important role in the lipid uptake determining the ligand specificity. Our results confirmed the previous assumption regarding the membrane-mediated antimicrobial action of Lc-LTP2 and afforded molecular insight into its biological role in the plant.

  2. Ovchinnikova T.V., Finkina E.I., Melnikova D.N., Bogdanov I.V. (2016). Plant pathogenesis-related proteins PR-10 and PR-14 as components of innate immunity system and ubiquitous allergens. Curr. Med. Chem. , [+]

    Pathogenesis-related (PR) proteins are components of innate immunity system in plants. They play an important role in plant defense against pathogens. Lipid transfer proteins (LTPs) and Bet v 1 homologues comprise two separate families of PR-proteins. Both LTPs (PR-14) and Bet v 1 homologues (PR-10) are multifunctional small proteins involving in plant response to abiotic and biotic stress conditions. The representatives of these PR-protein families do not show any sequence similarity but have other common biochemical features such as low molecular masses, the presence of hydrophobic cavities, ligand binding properties, and antimicrobial activities. Besides, many members of PR-10 and PR-14 families are ubiquitous plant panallergens which are able to cause sensitization of human immune system and cross-reactive allergic reactions to plant food and pollen. This review is aimed at comparative analysis of structure-functional and allergenic properties of the PR-10 and PR-14 families, as well as prospects for their medicinal application.

  3. Bogdanov I.V., Shenkarev Z.O., Finkina E.I., Melnikova D.N., Rumynskiy E.I., Arseniev A.S., Ovchinnikova T.V. (2016). A novel lipid transfer protein from the pea Pisum sativum: isolation, recombinant expression, solution structure, antifungal activity, lipid binding, and allergenic properties. BMC Plant Biol. 16 (1), 107 [+]

    Plant lipid transfer proteins (LTPs) assemble a family of small (7-9 kDa) ubiquitous cationic proteins with an ability to bind and transport lipids as well as participate in various physiological processes including defense against phytopathogens. They also form one of the most clinically relevant classes of plant allergens. Nothing is known to date about correlation between lipid-binding and IgE-binding properties of LTPs. The garden pea Pisum sativum is widely consumed crop and important allergenic specie of the legume family. This work is aimed at isolation of a novel LTP from pea seeds and characterization of its structural, functional, and allergenic properties.

  4. Finkina E.I., Melnikova D.N., Bogdanov I.V., Ovchinnikova T.V. (2016). Lipid Transfer Proteins As Components of the Plant Innate Immune System: Structure, Functions, and Applications. Acta Naturae 8 (2), 47–61 [+]

    Among a variety of molecular factors of the plant innate immune system, small proteins that transfer lipids and exhibit a broad spectrum of biological activities are of particular interest. These are lipid transfer proteins (LTPs). LTPs are interesting to researchers for three main features. The first feature is the ability of plant LTPs to bind and transfer lipids, whereby these proteins got their name and were combined into one class. The second feature is that LTPs are defense proteins that are components of plant innate immunity. The third feature is that LTPs constitute one of the most clinically important classes of plant allergens. In this review, we summarize the available data on the plant LTP structure, biological properties, diversity of functions, mechanisms of action, and practical applications, emphasizing their role in plant physiology and their significance in human life.

  5. Melnikova D.N., Mineev K.S., Finkina E.I., Arseniev A.S., Ovchinnikova T.V. (2016). A novel lipid transfer protein from the dill Anethum graveolens L.: isolation, structure, heterologous expression, and functional characteristics. J. Pept. Sci. 22 (1), 59–66 [+]

    A novel lipid transfer protein, designated as Ag-LTP, was isolated from aerial parts of the dill Anethum graveolens L. Structural, antimicrobial, and lipid binding properties of the protein were studied. Complete amino acid sequence of Ag-LTP was determined. The protein has molecular mass of 9524.4 Da, consists of 93 amino acid residues including eight cysteines forming four disulfide bonds. The recombinant Ag-LTP was overexpressed in Escherichia coli and purified. NMR investigation shows that the Ag-LTP spatial structure contains four α-helices, forming the internal hydrophobic cavity, and a long C-terminal tail. The measured volume of the Ag-LTP hydrophobic cavity is equal to ~800 A(3) , which is much larger than those of other plant LTP1s. Ag-LTP has weak antifungal activity and unpronounced lipid binding specificity but effectively binds plant hormone jasmonic acid. Our results afford further molecular insight into biological functions of LTP in plants. Copyright © 2015 European Peptide Society and John Wiley & Sons, Ltd.

  6. Алексеева А.С., Третьякова Д.С., Мельникова Д.Н., Молотковский Юл.Г., Болдырев И.А. (2016). Новый флуоресцентный мембранный зонд (2,3;5,6 бисциклогексил)bodipy меченный фосфатидилхолн. Биоорг. хим. 42 (3), 339–344 [+]

    Новый мембранный зонд бисциклогексил BODIPY (BCHB) меченный фосфатидилхолин структурно очень близок в 1,3,5,7 тетраметил BODIPY (TMB) меченному фосфатидилхолину и синтезируется по аналогичной схеме. Системы сопряженных связей BCHB и TMB формально идентичны, однако спектральные характеристики BCHB заметно отличаются, что делают BCHB хорошим акцептором фёрстеровского резонансного переноса (FRET) для TMB. Показано, что FRET пара фосфатидилхолинов на основе BCHB и TMB является перспективным инструментом для исследования мембранных систем, например, межмембранного липидного переноса.

  7. Bogdanov I.V., Finkina E.I., Balandin S.V., Melnikova D.N., Stukacheva E.A., Ovchinnikova T.V. (2015). Structural and Functional Characterization of Recombinant Isoforms of the Lentil Lipid Transfer Protein. Acta Naturae 7 (3), 65–73 [+]

    The recombinant isoforms Lc-LTP1 and Lc-LTP3 of the lentil lipid transfer protein were overexpressed in E. coli cells. It was confirmed that both proteins are stabilized by four disulfide bonds and characterized by a high proportion of the α-helical structure. It was found that Lc-LTP1 and Lc-LTP3 possess antimicrobial activity and can bind fatty acids. Both isoforms have the ability to bind specific IgE from sera of patients with food allergies, which recognize similar epitopes of the major peach allergen Pru p 3. Both isoforms were shown to have immunological properties similar to those of other plant allergenic LTPs, but Lc-LTP3 displayed a less pronounced immunoreactivity.

  8. Gizatullina A.K., Finkina E.I., Mineev K.S., Melnikova D.N., Bogdanov I.V., Telezhinskaya I.N., Balandin S.V., Shenkarev Z.O., Arseniev A.S., Ovchinnikova T.V. (2013). Recombinant production and solution structure of lipid transfer protein from lentil Lens culinaris. Biochem. Biophys. Res. Commun. 439 (4), 427–32 [+]

    Lipid transfer protein, designated as Lc-LTP2, was isolated from seeds of the lentil Lens culinaris. The protein has molecular mass 9282.7Da, consists of 93 amino acid residues including 8 cysteines forming 4 disulfide bonds. Lc-LTP2 and its stable isotope labeled analogues were overexpressed in Escherichia coli and purified. Antimicrobial activity of the recombinant protein was examined, and its spatial structure was studied by NMR spectroscopy. The polypeptide chain of Lc-LTP2 forms four α-helices (Cys4-Leu18, Pro26-Ala37, Thr42-Ala56, Thr64-Lys73) and a long C-terminal tail without regular secondary structure. Side chains of the hydrophobic residues form a relatively large internal tunnel-like lipid-binding cavity (van der Waals volume comes up to ∼600Å(3)). The side-chains of Arg45, Pro79, and Tyr80 are located near an assumed mouth of the cavity. Titration with dimyristoyl phosphatidylglycerol (DMPG) revealed formation of the Lc-LTP2/lipid non-covalent complex accompanied by rearrangements in the protein spatial structure and expansion of the internal cavity. The resultant Lc-LTP2/DMPG complex demonstrates limited lifetime and dissociates within tens of hours.