Group of in silico analysis of membrane proteins structure

NamePositionContacts
Anton Chugunov, Ph.D.headbatch2k@yandex.ru+7(915)1088825
Irina Paninaj. r. f.irinaspanina@gmail.com

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Anton Chugunov

  • Russia, Moscow, Ul. Miklukho-Maklaya 16/10 — On the map
  • IBCh RAS, build. 31, office. 608
  • Phone: +7(915)1088825
  • E-mail: batch2k@yandex.ru

Key factors contributing to the green-to-red fluorescent protein transformation were identified

In collaboration with Laboratory of biomolecular modeling,  Laboratory of molecular theranostics

Through the examples of two highly homologous fluorescent proteins from Zoanthus sp. (zoanGFP and zoan2RFP), amino acid residues participating in the transformation of a protein with the green fluorescence (GFP) into the red fluorescent protein (RFP) were explored. As the result of zoanGFP mutagenesis, internal amino acid residues (a.a.r.) became identical to those of zoan2RFP. However, this mutant underwent only partial transformation into the red form. To elucidate the extra factors that might affect red chromophore biosynthesis, we used comparative molecular dynamics simulations of zoan2RFP and zoanGFPmut. As the result, additional a.a.r. were discovered on the surface of the protein that might influence both the arrangement and flexibility of the chromophore-surrounding a.a.r. Site-directed mutagenesis of these external a.a.r. confirmed the crucial role of these residues in red chromophore biosynthesis.

Combinatorial selective incorporation of stable13C and 15N isotopes facilitates NMR spectra analysis and allows mapping of the binding interfaces between membrane receptors and their ligands

In collaboration with Laboratory of bioengineering of neuromodulators and neuroreceptors,  Laboratory of Molecular Instruments for Neurobiology,  Laboratory of structural biology of ion channels

Combinatorial incorporation of stable 13C and 15N isotopes into protein molecules can significantly simplify the analysis of NMR spectra. For the first time, the problem was solved and the CombLabel algorithm was developed for calculating combinatorial 13C and 15N  labeling schemes with a minimum price. The application of the program allowed to assign 50% of the NMR signals of the backbone of the second voltage-sensing domain of human sodium channel Nav1.4 (VSD-II). Leak currents through mutant variants of Nav1.4 containing Arg675Gly mutation in VSD-II lead to the development of a hereditary disease – normokalemic periodic paralysis. Hm-3 toxin from the venom of spider Heriaeus melloteei is able to block leak currents in VSD-II. By the means of NMR spectroscopy the interaction interface between VSD-II and Hm-3 toxin was determined. According to the model of the VSD-II/Hm-3 complex, based on the NMR data, the toxin binds to the extracellular S1-S2 loop, destabilizing the state of the domain, at which leak currents are observed. Using the example of the complexes of Hm-3 toxin with VSD-I and VSD-II of the Nav1.4 channel, it has been shown that arachnid toxins can interact differently with different domains within the same sodium channel.

Protein Surface Topography was used to improve a potassium channel blocker

In collaboration with Laboratory of biomolecular NMR-spectroscopy,  Laboratory of biomolecular modeling,  Laboratory of Molecular Instruments for Neurobiology

Previously, for the design of peptides with a given function, we have proposed using a convenient structural framework, namely, the α-hairpinin fold, characteristic of toxins from scorpion venom and plant defense peptides. Now, the use of the Protein Surface Topography method that we developed, has significantly improved the properties of an artificial α-hairpinin, which blocks Kv1.3 potassium channels, an important pharmacological target. The joint application of two approaches, namely, scaffold engineering and protein surface topography, can be used to obtain optimized ion channel ligands.

KV1.2 Сhannel-Specific Blocker from Scorpion Venom: Structural Basis of Selectivity

In collaboration with Laboratory of optical microscopy and spectroscopy of biomolecules,  Group of nanobioengineering,  Laboratory of Molecular Instruments for Neurobiology

Laboratory of Molecular Instruments for Neurobiology is known for systematic study of Arthropods’ venoms and derived peptides that specifically target various ion channels. Scorpions’ venom is abundant with potassium channels (Kv) blockers, and this diversity was described in previously released in Kalium database.

In cooperation with Laboratory of optical microscopy and spectroscopy of biomolecules and Group of nanobioengineering an unique screening system permitted identification in the Mesobuthus eupeus scorpion venom of Kv1.2 blocker: peptide MeKTx11-1 binging with high affinity (IC50 ≈0,2 nM) and specificity (effect on Kv1.1, 1.3 and 1.6 emerges at >100-fold higher concentrations). This peptide differs from the related MeKTx11-3 by just two residues, possessing substantially lower Kv1.2-specificity.

Finally, Group of in silico analysis of membrane proteins structure conducted a molecular modeling study of these two peptides interacting with Kv1.2 channel, immersed into an explicit lipid bilayer. This study uncovered mechanism of selective action of MeKTx11-1 peptide. The developed analysis technique will be of use for future design of selective ligands of Kv and other channels, which may be applied in fundamental studies of molecular basis of nervous system function and as drugs prototypes.

MeKTx11-1, Kv1.2 channel –specific peptide blocker from the M.eupeus scorpion venom: structural basis of selectivity

In collaboration with Laboratory of biomolecular modeling,  Group of nanobioengineering,  Laboratory of Molecular Instruments for Neurobiology,  Laboratory of optical microscopy and spectroscopy of biomolecules

A.V. Feofanov (Laboratory of optical microscopy and spectroscopy of biomolecules), О.V. Nekrasova, K.S.Kudryashova (Group of nanobioengineering, Bioengineering department), A.A. Vassilevski, A.I. Kuzmenkov, A.M. Gigolaev (Laboratory of molecular instruments for neurobiology), A.O. Chugunov, V.M. Tabakmakher, R.G. Efremov (Group of in silico analysis of membrane proteins structure, Laboratory of biomolecular modeling).

A unique high-affinity and highly selective peptide blocker of Kv1.2 channel, MeKTx11-1, from the scorpion venom Mesobuthus eupeus was studied. Peptide MeKTx11-1 and its mutant forms were produced in a recombinant form, and their receptor-binding activity was studied against a panel of Kv1-channels. Molecular modeling of interaction of these peptides with Kv1.2 channel was carried out, and key structural elements of the interactions were determined. Peptide MeKTx11-1 may be used as a novel efficient molecular tool in neurobiology to identify and study the activity of Kv1.2 channel in the presence of different isoforms of Kv1-channels.

In collaboration with S.Peigneur and J.Tytgat fromUniversity of Leuven, Belgium and A.F. Fradkov from Evrogen JSC.

Scorpion venom is rich in peptide blockers of voltage-gated potassium channels (KV), and we have reflected this diversity previously in Kalium, a database dedicated to such peptides. A high-affinity and selective blocker of KV1.2 channels, characteristic of the human central nervous system, was obtained from the venom of the scorpion Mesobuthus eupeus. Using molecular modeling and site-directed mutagenesis, the mechanism of selective interaction between the toxin and channels was investigated.

MeKTx11-1, Kv1.2 channel –specific peptide blocker from the M.eupeus scorpion venom: structural basis of selectivity

In collaboration with Laboratory of optical microscopy and spectroscopy of biomolecules,  Laboratory of Molecular Instruments for Neurobiology,  Group of nanobioengineering

Оksana V. Nekrasova, K.S.Kudryashova (Group of nanobioengineering, Bioengineering department), A.A. Vassilevski, A.I. Kuzmenkov, A.M. Gigolaev (Laboratory of molecular instruments for neurobiology), A.O. Chugunov, V.M. Tabakmakher, R.G. Efremov (Group of in silico analysis of membrane proteins structure, Laboratory of biomolecular modeling), A.V. Feofanov (Laboratory of optical microscopy and spectroscopy of biomolecules).

A unique high-affinity and highly selective peptide blocker of Kv1.2 channel, MeKTx11-1, from the scorpion venom Mesobuthus eupeus was studied. Peptide MeKTx11-1 and its mutant forms were produced in a recombinant form, and their receptor-binding activity was studied against a panel of Kv1-channels. Molecular modeling of interaction of these peptides with Kv1.2 channel was carried out, and key structural elements of the interactions were determined. Peptide MeKTx11-1 may be used as a novel efficient molecular tool in neurobiology to identify and study the activity of Kv1.2 channel in the presence of different isoforms of Kv1-channels.

In collaboration with S.Peigneur and J.Tytgat fromUniversity of Leuven, Belgium and A.F. Fradkov from Evrogen JSC.