Novel Thionins from Black Seed (Nigella sativa L.) Demonstrate Antimicrobial Activity
- 248 Downloads
Black seed (Nigella sativa) is known as a good source of various biologically active compounds which possess antimicrobial properties. One of our objectives was to elaborate methods of obtaining and extracting peptides from plants. In the current study, we discovered some biological effects of thionins from black seed, such as bactericidal and fungicidal effects. Isolation of thionins performed by combining acidic extraction and fractionation with various liquid chromatography methods. The N-terminal amino acid sequences were revealed using automated Edman degradation. The antimicrobial activity of thionins were evaluated by a microdilution broth assay. A fluorescent spectroscopy and an atomic force microscopy allow to investigate the features of mode of action of the thionins. The two novel peptides from black seed (N. sativa L.), a plant endemic to Central Asia. These peptides, named NsW1 and NsW2, have a high affinity with heparin, a polysaccharide glycosaminoglycan. These molecules were indentified as thionins, a well-known family of plant antimicrobial peptides. These thionins effectively inhibit viability of Bacillus subtilus, Staphylococcus aureus and Candida albicans that has been confirmed using a bacteriological and some biophysical techniques. Obtained data indicate that black seed thionins are biologically active molecules that may be considered to be perspective antibacterial agents.
KeywordsBlack seed Thionins Antimicrobial activity Atomic force microscopy
This work was supported by Russian Scientific Foundation (Grant No. 14-50-00131) (section devoted of “Isolation and srtuctural characterization of the black seed thionins”), Russian foundation of Basic Research (Grant No. 16-34-60217-mol_a_dk) (section devoted of “Antimicrobial and cytotoxic activity of the black seed thionins”) and Stipend of The President of Russian Federation for young scientists (Registration No. SP-943.2015.4, SP- 2093.2015.4). The authors are grateful to Yulia I. Oshchepkova and Dr. Olga N. Veshkurova (both from Laboratory of Protein and Peptide Chemistry A.S. Sadykov Institute of Bioorganic Chemistry Academy of Sciences the Republic of Uzbekistan) for introduced defatted seeds of N. sativa and the initial stages of the seed extract fractionation; Dr. Elena V. Svirshchevskaya (from Laboratory of Cellular Interactions Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry Russians Academy of Science) for cytotoxity assays of the black seed thionins; Prof. G.D. Zasukhina (from Vavilov Institute of General Genetics Russian Academy of Sciences) and Dr. V.F. Mikhailov (from Burnasyan Federal Medical Biophysical Center of FMBA of Russia) for the important suggestions and advices while the manuscript was in progress.
Compliance with Ethical Standards
Conflict of interest
Alexey S. Vasilchenko, Alexey N. Smirnov, Sergei K. Zavriev, Eugene V. Grishin, Anastasia V. Vasilchenko and Eugene A. Rogozhin confirm that this article content has no conflicts of interest.
Human and Animal Rights
This article does not contain studies with human or animal subjects performed by any of the authors that should be approved by Ethics Committee.
The article does not contain any studies in patients by any of the authors.
- Bohlmann H, Clausen S, Behnke S, Giese H, Hiller C, Reimann-Philipp U, Schrader G, Barkholt V, Apel K (1988) Leaf-specific thionins of barley-a novel class of cell wall proteins toxic to plant-pathogenic fungi and possibly involved in the defence mechanism of plants. EMBO J 7:1559–1565PubMedPubMedCentralGoogle Scholar
- Debreczen JE, Girmann B, Zeeck A, Kratzner R, Sheldrick GM (2003) Structure of viscotoxin A3: disulfide location from weak SAD data. Acta Cryst 59:2125–2132Google Scholar
- Li L, Sun J, Xia S, Tian X, Cheserek MJ, Le G (2016) Mechanism of antifungal activity of antimicrobial peptide APP, a cell-penetrating peptide derivative, against Candida albicans: intracellular DNA binding and cell cycle arrest. Appl Microbiol Biotechnol 100(7):3245–3253. doi: 10.1007/s00253-015-7265-y CrossRefPubMedGoogle Scholar
- Nolde SB, Vassilevski AA, Rogozhin EA, Barinov NA, Balashova TA, Samsonova OV, Baranov YV, Feofanov AV, Egorov TA, Arseniev AS, Grishin EV (2011) Disulfide-stabilized helical hairpin structure of a novel antifungal peptide EcAMP1 from seeds of barnyard grass (Echinochloa crus-galli). J Biol Chem 286:25145–25153CrossRefPubMedPubMedCentralGoogle Scholar
- Slavokhotova AA, Rogozhin EA, Musolyamov AK, Andreev YA, Oparin PB, Berkut AA, Vassilevski AA, Egorov TA, Grishin EV, Odintsova TI (2014) Novel antifungal α-hairpinin peptide from Stellaria media seeds: structure, biosynthesis, gene structure and evolution. Plant Mol Biol 84:189–202CrossRefPubMedGoogle Scholar
- Stec B, Rao U, Teeter MM (1995) Refinement of purothionins reveals the solute particles important for the lattice formation and toxicity. Part 1: structure of β-putothionin at 1.7Å resolution. Acta Cryst 51:914–924Google Scholar
- Stuart LS, Harris TH (1942) Bactericidal and fungicidal properties of a crystalline protein from unbleached wheat flour. Cereal Chem 19:288–300Google Scholar
- Utkina LL, Andreev YA, Rogozhin EA, Korostyleva TV, Slavokhotova AA, Oparin PB, Vassilevski AA, Grishin EV, Egorov TA, Odintsova TI (2013) Genes encoding 4-Cys antimicrobial peptides in wheat Triticum kiharae Dorof. et Migush.: multimodular structural organization, instraspecific variability, distribution and role in defense. FEBS J 280:3594–3608CrossRefPubMedGoogle Scholar