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Your search keyword '"Vladislav A. Lushpa"' showing total 4 results
Start Over Author "Vladislav A. Lushpa" Topic medicine
4 results on '"Vladislav A. Lushpa"'

1. Peptides from the Sea Anemone Metridium senile with Modified Inhibitor Cystine Knot (ICK) Fold Inhibit Nicotinic Acetylcholine Receptors

Author

Igor E. Kasheverov, Yulia A. Logashina, Fedor D. Kornilov, Vladislav A. Lushpa, Ekaterina E. Maleeva, Yuliya V. Korolkova, Jinpeng Yu, Xiaopeng Zhu, Dongting Zhangsun, Sulan Luo, Klara Stensvåg, Denis S. Kudryavtsev, Konstantin S. Mineev, and Yaroslav A. Andreev

Subjects
sea anemone venom, Metridium senile, ICK fold peptides, nicotinic acetylcholine receptors, NMR structure, electrophysiology, Medicine
Abstract

Nicotinic acetylcholine receptors (nAChRs) play an important role in the functioning of the central and peripheral nervous systems, and other organs of living creatures. There are several subtypes of nAChRs, and almost all of them are considered as pharmacological targets in different pathological states. The crude venom of the sea anemone Metridium senile showed the ability to interact with nAChRs. Four novel peptides (Ms11a-1–Ms11a-4) with nAChR binding activity were isolated. These peptides stabilized by three disulfide bridges have no noticeable homology with any known peptides. Ms11a-1–Ms11a-4 showed different binding activity towards the muscle-type nAChR from the Torpedo californica ray. The study of functional activity and selectivity for the most potent peptide (Ms11a-3) revealed the highest antagonism towards the heterologous rat α9α10 nAChR compared to the muscle and α7 receptors. Structural NMR analysis of two toxins (Ms11a-2 and Ms11a-3) showed that they belong to a new variant of the inhibitor cystine knot (ICK) fold but have a prolonged loop between the fifth and sixth cysteine residues. Peptides Ms11a-1–Ms11a-4 could represent new pharmacological tools since they have structures different from other known nAChRs inhibitors.

Published
2022
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2. Modulation of Toll-like receptor 1 intracellular domain structure and activity by Zn2+ ions

Author

Daniil Vakhrameev, Xiaohui Wang, Irina A. Talyzina, Arthur O. Zalevsky, Sergey A. Goncharuk, Mikhail B. Shevtsov, Konstantin S. Mineev, Valentin Borshchevskiy, Vladislav A. Lushpa, Alexander S. Arseniev, Marina V. Goncharuk, Aleksandra Luginina, and Cong Lin

Subjects
QH301-705.5, In silico, Medicine (miscellaneous), medicine.disease_cause, digestive system, General Biochemistry, Genetics and Molecular Biology, Article, Protein Domains, ddc:570, parasitic diseases, Extracellular, medicine, Humans, Biology (General), Receptor, X-ray crystallography, Ions, Toll-like receptor, Mutation, Chemistry, biochemical phenomena, metabolism, and nutrition, Toll-Like Receptor 1, Toll-Like Receptor 2, Toll-like receptors, Zinc, HEK293 Cells, Cytoplasm, Biophysics, Molecular modelling, General Agricultural and Biological Sciences, Solution-state NMR, Intracellular, Cysteine
Abstract

Toll-like receptors (TLRs) play an important role in the innate immune response. While a lot is known about the structures of their extracellular parts, many questions are still left unanswered, when the structural basis of TLR activation is analyzed for the TLR intracellular domains. Here we report the structure and dynamics of TLR1 toll-interleukin like (TIR) cytoplasmic domain in crystal and in solution. We found that the TLR1-TIR domain is capable of specific binding of Zn with nanomolar affinity. Interactions with Zn are mediated by cysteine residues 667 and 686 and C667 is essential for the Zn binding. Potential structures of the TLR1-TIR/Zn complex were predicted in silico. Using the functional assays for the heterodimeric TLR1/2 receptor, we found that both Zn addition and Zn depletion affect the activity of TLR1, and C667A mutation disrupts the receptor activity. Analysis of C667 position in the TLR1 structure and possible effects of C667A mutation, suggests that zinc-binding ability of TLR1-TIR domain is critical for the receptor activation., Lushpa et al report the structure and dynamics of the TLR1 toll-interleukin like (TIR) cytoplasmic domain in both crystal and solution. They demonstrate that the TLR1 TIR domain is capable of specific binding of Zn with nanomolar affinity, which appears to be critical for receptor activation, and provide potential structures TLR1-TIR/Zn complex based on in silico data.

Published
2021

3. Sampling the cultivation parameter space for the bacterial production of TLR1 intracellular domain reveals the multiple optima

Author

Sergey A. Goncharuk, Vladislav A. Lushpa, Konstantin S. Mineev, Marina V. Goncharuk, and Alexander S. Arseniev

Subjects
0106 biological sciences, Intracellular domain, Lysis, Gene Expression, Parameter space, medicine.disease_cause, 01 natural sciences, 03 medical and health sciences, chemistry.chemical_compound, Protein Domains, 010608 biotechnology, Escherichia coli, medicine, Humans, 030304 developmental biology, 0303 health sciences, Chemistry, Toll-Like Receptor 1, Recombinant Proteins, Yield (chemistry), Triton X-100, Biophysics, Protein solubility, Intracellular, Biotechnology
Abstract

T7 expression system is an extremely popular approach for the recombinant protein production in Escherichia coli for structural and functional studies and therapeutic applications. There are many useful tools and successful techniques that allow expressing the desired protein in this system. However, high yield of soluble protein often requires a systematic optimization of a wide range of cell cultivation parameters. Here we analyze the effect of three key cultivation parameters - chemical inductor, temperature and time of post-induction culturing on the expression level of TLR1 intracellular TIR domain in a soluble form. In addition, the influence of Triton X-100 detergent on the protein solubility during the cell lysis was investigated. We show that a high expression level of the correctly folded soluble protein can be obtained under different combinations of cultivation parameters.

Published
2021

4. TIR domains of TLR family-from the cell culture to the protein sample for structural studies.

Author

Vladislav A Lushpa, Marina V Goncharuk, Irina A Talyzina, Alexander S Arseniev, Eduard V Bocharov, Konstantin S Mineev, and Sergey A Goncharuk

Subjects
Medicine, Science
Abstract

Toll-like receptors (TLRs) are key players in the innate immune system. Despite the great efforts in TLR structural biology, today we know the spatial structures of only four human TLR intracellular TIR domains. All of them belong to one of five subfamilies of receptors. One of the main bottlenecks is the high-level production of correctly folded proteins in soluble form. Here we used a rational approach to find the optimal parameters to produce TIR domains of all ten human TLR family members in soluble form in E. coli cells. We showed that dozens of milligrams of soluble His-tagged TLR2/3/6/7TIR and MBP-tagged TLR3/5/7/8TIR can be produced. We also developed the purification protocols and demonstrated by CD and NMR spectroscopy that purified TLR2/3/7TIR demonstrate a structural organization inherent to TIR domains. This illustrates the correct folding of produced proteins and their suitability for further structural and functional investigations.

Published
2024
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