Publications by authors named "Olga M Selivanova"

22 Publications

  • Page 1 of 1

New Model for Stacking Monomers in Filamentous Actin from Skeletal Muscles of .

Int J Mol Sci 2020 Nov 6;21(21). Epub 2020 Nov 6.

Institute of Protein Research, Russian Academy of Sciences, 142290 Pushchino, Moscow Region, Russia.

To date, some scientific evidence (limited proteolysis, mass spectrometry analysis, electron microscopy (EM)) has accumulated, which indicates that the generally accepted model of double-stranded of filamentous actin (F-actin) organization in eukaryotic cells is not the only one. This entails an ambiguous understanding of many of the key cellular processes in which F-actin is involved. For a detailed understanding of the mechanism of F-actin assembly and actin interaction with its partners, it is necessary to take into account the polymorphism of the structural organization of F-actin at the molecular level. Using electron microscopy, limited proteolysis, mass spectrometry, X-ray diffraction, and structural modeling we demonstrated that F-actin presented in the EM images has no double-stranded organization, the regions of protease resistance are accessible for action of proteases in F-actin models. Based on all data, a new spatial model of filamentous actin is proposed, and the F-actin polymorphism is discussed.
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http://dx.doi.org/10.3390/ijms21218319DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7664232PMC
November 2020

Antimicrobial and Amyloidogenic Activity of Peptides Synthesized on the Basis of the Ribosomal S1 Protein from Thermus Thermophilus.

Int J Mol Sci 2020 09 2;21(17). Epub 2020 Sep 2.

Institute of Protein Research, Russian Academy of Sciences, 142290 Pushchino, Moscow Region, Russia.

Controlling the aggregation of vital bacterial proteins could be one of the new research directions and form the basis for the search and development of antibacterial drugs with targeted action. Such approach may be considered as an alternative one to antibiotics. Amyloidogenic regions can, like antibacterial peptides, interact with the "parent" protein, for example, ribosomal S1 protein (specific only for bacteria), and interfere with its functioning. The aim of the work was to search for peptides based on the ribosomal S1 protein from , exhibiting both aggregation and antibacterial properties. The biological system of the response of Gram-negative bacteria to the action of peptides was characterized. Among the seven studied peptides, designed based on the S1 protein sequence, the R23I (modified by the addition of HIV transcription factor fragment for bacterial cell penetration), R23T (modified), and V10I (unmodified) peptides have biological activity that inhibits the growth of cells, that is, they have antimicrobial activity. But, only the R23I peptide had the most pronounced activity comparable with the commercial antibiotics. We have compared the proteome of peptide-treated and intact cells. These important data indicate a decrease in the level of energy metabolism and anabolic processes, including the processes of biosynthesis of proteins and nucleic acids. Under the action of 20 and 50 μg/mL R23I, a decrease in the number of proteins in cells was observed and S1 ribosomal protein was absent. The obtained results are important for understanding the mechanism of amyloidogenic peptides with antimicrobial activity and can be used to develop new and improved analogues.
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http://dx.doi.org/10.3390/ijms21176382DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7504387PMC
September 2020

Amyloidogenic Propensities of Ribosomal S1 Proteins: Bioinformatics Screening and Experimental Checking.

Int J Mol Sci 2020 Jul 22;21(15). Epub 2020 Jul 22.

Institute of Protein Research, Russian Academy of Sciences, Pushchino 142290, Moscow Region, Russia.

Structural S1 domains belong to the superfamily of oligosaccharide/oligonucleotide-binding fold domains, which are highly conserved from prokaryotes to higher eukaryotes and able to function in RNA binding. An important feature of this family is the presence of several copies of the structural domain, the number of which is determined in a strictly limited range from one to six. Despite the strong tendency for the aggregation of several amyloidogenic regions in the family of the ribosomal S1 proteins, their fibril formation process is still poorly understood. Here, we combined computational and experimental approaches for studying some features of the amyloidogenic regions in this protein family. The FoldAmyloid, Waltz, PASTA 2.0 and Aggrescan programs were used to assess the amyloidogenic propensities in the ribosomal S1 proteins and to identify such regions in various structural domains. The thioflavin T fluorescence assay and electron microscopy were used to check the chosen amyloidogenic peptides' ability to form fibrils. The bioinformatics tools were used to study the amyloidogenic propensities in 1331 ribosomal S1 proteins. We found that amyloidogenicity decreases with increasing sizes of proteins. Inside one domain, the amyloidogenicity is higher in the terminal parts. We selected and synthesized 11 amyloidogenic peptides from the and ribosomal S1 proteins and checked their ability to form amyloids using the thioflavin T fluorescence assay and electron microscopy. All 11 amyloidogenic peptides form amyloid-like fibrils. The described specific amyloidogenic regions are actually responsible for the fibrillogenesis process and may be potential targets for modulating the amyloid properties of bacterial ribosomal S1 proteins.
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http://dx.doi.org/10.3390/ijms21155199DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7432502PMC
July 2020

Structural and Functional Peculiarities of α-Crystallin.

Biology (Basel) 2020 Apr 23;9(4). Epub 2020 Apr 23.

Institute of Protein Research, Russian Academy of Sciences, 142290 Pushchino, Moscow Region, Russia.

α-Crystallin is the major protein of the eye lens and a member of the family of small heat-shock proteins. Its concentration in the human eye lens is extremely high (about 450 mg/mL). Three-dimensional structure of native α-crystallin is unknown. First of all, this is the result of the highly heterogeneous nature of α-crystallin, which hampers obtaining it in a crystalline form. The modeling based on the electron microscopy (EM) analysis of α-crystallin preparations shows that the main population of the α-crystallin polydisperse complex is represented by oligomeric particles of rounded, slightly ellipsoidal shape with the diameter of about 13.5 nm. These complexes have molecular mass of about 700 kDa. In our opinion, the heterogeneity of the α-crystallin complex makes it impossible to obtain a reliable 3D model. In the literature, there is evidence of an enhanced chaperone function of α-crystallin during its dissociation into smaller components. This may indirectly indicate that the formation of heterogeneous complexes is probably necessary to preserve α-crystallin in a state inactive before stressful conditions. Then, not only the heterogeneity of the α-crystallin complex is an evolutionary adaptation that protects α-crystallin from crystallization but also the enhancement of the function of α-crystallin during its dissociation is also an evolutionary acquisition. An analysis of the literature on the study of α-crystallin in vitro led us to the assumption that, of the two α-crystallin isoforms (αA- and αB-crystallins), it is αA-crystallin that plays the role of a special chaperone for αB-crystallin. In addition, our data on X-ray diffraction analysis of α-crystallin at the sample concentration of about 170-190 mg/mL allowed us to assume that, at a high concentration, the eye lens α-crystallin can be in a gel-like stage. Finally, we conclude that, since all the accumulated data on structural-functional studies of α-crystallin were carried out under conditions far from native, they cannot adequately reflect the features of the functioning of α-crystallin in vivo.
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http://dx.doi.org/10.3390/biology9040085DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7235859PMC
April 2020

The number of domains in the ribosomal protein S1 as a hallmark of the phylogenetic grouping of bacteria.

PLoS One 2019 22;14(8):e0221370. Epub 2019 Aug 22.

Institute of Protein Research, Russian Academy of Sciences, Pushchino, Moscow Region, Russia.

The family of ribosomal proteins S1 contains about 20% of all bacterial proteins including the S1 domain. An important feature of this family is multiple copies of structural domains in bacteria, the number of which changes in a strictly limited range from one to six. In this study, the automated exhaustive analysis of 1453 sequences of S1 allowed us to demonstrate that the number of domains in S1 is a distinctive characteristic for phylogenetic bacterial grouping in main phyla. 1453 sequences of S1 were identified in 25 out of 30 different phyla according to the List of Prokaryotic Names with Standing in Nomenclature. About 62% of all records are identified as six-domain S1 proteins, which belong to phylum Proteobacteria. Four-domain S1 are identified mainly in proteins from phylum Firmicutes and Actinobacteria. Records belonging to these phyla are 33% of all records. The least represented two-domain S1 are about 0.6% of all records. The third and fourth domains for the most representative four- and six-domain S1 have the highest percentage of identity with the S1 domain from polynucleotide phosphorylase and S1 domains from one-domain S1. In addition, for these groups, the central part of S1 (the third domain) is more conserved than the terminal domains.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0221370PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6705787PMC
March 2020

Should the Treatment of Amyloidosis Be Personified? Molecular Mechanism of Amyloid Formation by Aβ Peptide and Its Fragments.

J Alzheimers Dis Rep 2018 Oct 24;2(1):181-199. Epub 2018 Oct 24.

Institute of Protein Research, Russian Academy of Science, Pushchino, Moscow Region, Russia.

Aβ and Aβ peptides are believed to be associated with Alzheimer's disease. Aggregates (plaques) of Aβ fibrils are found in the brains of humans affected with this disease. The mechanism of formation of Aβ fibrils has not been studied completely, which hinders the development of a correct strategy for therapeutic prevention of this neurodegenerative disorder. It has been found that the most toxic samples upon generation of fibrils are different oligomeric formations. Based on different research methods used for studying amyloidogenesis of Aβ and Aβ peptides and its amyloidogenic fragments, we have proposed a new mechanism of formation of amyloid fibrils. In accord with this mechanism, the main building unit for fibril generation is a ring-like oligomer. Association of ring-like oligomers results in the formation of fibrils of different morphologies. Our model implies that to prevent development of Alzheimer's disease a therapeutic intervention is required at the earliest stages of amyloidogenesis-at the stage of formation of ring-like oligomers. Therefore, the possibility of a personified approach for prevention not only of Alzheimer's disease development but also of other neurodegenerative diseases associated with the formation of fibrils is argued.
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http://dx.doi.org/10.3233/ADR-180063DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6218156PMC
October 2018

To Be Fibrils or To Be Nanofilms? Oligomers Are Building Blocks for Fibril and Nanofilm Formation of Fragments of Aβ Peptide.

Langmuir 2018 02 30;34(6):2332-2343. Epub 2018 Jan 30.

Institute of Protein Research, Russian Academy of Sciences , Pushchino 142290, Russia.

To identify the key stages in the amyloid fibril formation we studied the aggregation of amyloidogenic fragments of Aβ peptide, Aβ(16-25), Aβ(31-40), and Aβ(33-42), using the methods of electron microscopy, X-ray analysis, mass spectrometry, and structural modeling. We have found that fragments Aβ(31-40) and Aβ(33-42) form amyloid fibrils in the shape of bundles and ribbons, while fragment Aβ(16-25) forms only nanofilms. We are the first who performed 2D reconstruction of amyloid fibrils by the Markham rotation technique on electron micrographs of negatively stained fragments of Aβ peptide. Combined analysis of the data allows us to speculate that both the fibrils and the films are formed via association of ring-shaped oligomers with the external diameter of about 6 to 7 nm, the internal diameter of 2 to 3 nm, and the height of ∼3 nm. We conclude that such oligomers are the main building blocks in fibrils of any morphology. The interaction of ring oligomers with each other in different ways makes it possible to explain their polymorphism. The new mechanism of polymerization of amyloidogenic proteins and peptides, described here, could stimulate new approaches in the development of future therapeutics for the treatment of amyloid-related diseases.
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http://dx.doi.org/10.1021/acs.langmuir.7b03393DOI Listing
February 2018

Rosetta Stone for Amyloid Fibrils: The Key Role of Ring-Like Oligomers in Amyloidogenesis.

J Alzheimers Dis 2017 ;59(3):785-795

Institute of Protein Research, Russian Academy of Sciences, Pushchino, Russia.

Deeper understanding of processes of protein misfolding, aggregation, formation of oligomers, protofibrils, and fibrils is crucial for the development of future medicine in treatment of amyloid-related diseases. While numerous reports illuminate the field, the above processes are extremely complex, as they depend on many varying parameters, such as the peptide concentration, temperature, pH, presence of metal ions, lipids, and organic solvents. Different mechanisms of amyloid fibril formation have been proposed, but the process of the oligomer-to-fibril transition is the least agreed upon. Our studies of a number of amyloidogenic proteins and peptides (insulin, Aβ peptides, the Bgl2 protein from the yeast cell wall), as well as their amyloidogenic fragments, have allowed us to propose a model of the fibril structure generation. We have found that the main building block of fibrils of any morphology is a ring-like oligomer. The varying models of interaction of ring oligomers with each other revealed in our studies make it possible to explain their polymorphism. Crucially, the amino acid sequence determines the oligomer structure for the given protein/peptide.
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http://dx.doi.org/10.3233/JAD-170230DOI Listing
April 2018

Different amyloid aggregation of smooth muscles titin in vitro.

J Biomol Struct Dyn 2018 Jul 10;36(9):2237-2248. Epub 2017 Jul 10.

a Institute of Theoretical and Experimental Biophysics, Russian Academy of Sciences , Pushchino , Moscow Region 142290 , Russia.

A comparative study of amyloid properties of the aggregates of smooth muscle titin (SMT) from chicken gizzard was carried out. These aggregates were formed in two solutions: 0.15 M glycine-KOH, pH 7.2-7.4 (SMT(Gly)) and 0.2 M KCl, 10 mM imidazole, pH 7.0 (SMT(KCl)). Electron microscopy data showed that SMT aggregates has an amorphous structure in both cases. The results of atomic-force microscopy demonstrated slight differences in morphology in two types of aggregates. The SMT(Gly) aggregates were represented as branching chains, composed of spherical aggregates approximately 300-500 nm in diameter and up to 35 nm in height. The SMT(KCl) aggregates formed sponge-like structures with strands of 8-10 nm in height. Structural analysis of SMT aggregates by X-ray diffraction revealed the presence of cross-β-sheet structure in the samples under study. In the presence of SMT(Gly) aggregates, thioflavine T fluorescence intensity was higher (~3-fold times) compared with that in the presence of SMT(KCl) aggregates. Congo red-stained SMT(Gly) aggregates had yellow to apple-green birefringence under polarized light, which was not observed for SMT(KCl) aggregates. Dynamic light scattering data showed the similar rate of aggregation for both types of aggregates, though SMT(KCl) aggregates were able to partially disaggregate under increased ionic strength of the solution. The ability of SMT to aggregation followed by disaggregation may be functionally significant in the cell.
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http://dx.doi.org/10.1080/07391102.2017.1348988DOI Listing
July 2018

Taxonomic distribution, repeats, and functions of the S1 domain-containing proteins as members of the OB-fold family.

Proteins 2017 04 3;85(4):602-613. Epub 2017 Feb 3.

Group of Bioinformatics, Institute of Protein Research, Russian Academy of Sciences, Pushchino, Moscow Region, 142290, Russia.

Proteins of the nucleic acid-binding proteins superfamily perform such functions as processing, transport, storage, stretching, translation, and degradation of RNA. It is one of the 16 superfamilies containing the OB-fold in protein structures. Here, we have analyzed the superfamily of nucleic acid-binding proteins (the number of sequences exceeds 200,000) and obtained that this superfamily prevalently consists of proteins containing the cold shock DNA-binding domain (ca. 131,000 protein sequences). Proteins containing the S1 domain compose 57% from the cold shock DNA-binding domain family. Furthermore, we have found that the S1 domain was identified mainly in the bacterial proteins (ca. 83%) compared to the eukaryotic and archaeal proteins, which are available in the UniProt database. We have found that the number of multiple repeats of S1 domain in the S1 domain-containing proteins depends on the taxonomic affiliation. All archaeal proteins contain one copy of the S1 domain, while the number of repeats in the eukaryotic proteins varies between 1 and 15 and correlates with the protein size. In the bacterial proteins, the number of repeats is no more than 6, regardless of the protein size. The large variation of the repeat number of S1 domain as one of the structural variants of the OB-fold is a distinctive feature of S1 domain-containing proteins. Proteins from the other families and superfamilies have either one OB-fold or change slightly the repeat numbers. On the whole, it can be supposed that the repeat number is a vital for multifunctional activity of the S1 domain-containing proteins. Proteins 2017; 85:602-613. © 2016 Wiley Periodicals, Inc.
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http://dx.doi.org/10.1002/prot.25237DOI Listing
April 2017

The Mechanism Underlying Amyloid Polymorphism is Opened for Alzheimer's Disease Amyloid-β Peptide.

J Alzheimers Dis 2016 09;54(2):821-30

Institute of Protein Research, Russian Academy of Sciences, Pushchino, Moscow Region, Russia.

It has been demonstrated using Aβ40 and Aβ42 recombinant and synthetic peptides that their fibrils are formed of complete oligomer ring structures. Such ring structures have a diameter of about 8-9 nm, an oligomer height of about 2- 4 nm, and an internal diameter of the ring of about 3-4 nm. Oligomers associate in a fibril in such a way that they interact with each other, overlapping slightly. There are differences in the packing of oligomers in fibrils of recombinant and synthetic Aβ peptides. The principal difference is in the degree of orderliness of ring-like oligomers that leads to generation of morphologically different fibrils. Most ordered association of ring-like structured oligomers is observed for a recombinant Aβ40 peptide. Less ordered fibrils are observed with the synthetic Aβ42 peptide. Fragments of fibrils the most protected from the action of proteases have been determined by tandem mass spectrometry. It was shown that unlike Aβ40, fibrils of Aβ42 are more protected, showing less ordered organization compared to that of Aβ40 fibrils. Thus, the mass spectrometry data agree with the electron microscopy data and structural models presented here.
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http://dx.doi.org/10.3233/JAD-160405DOI Listing
September 2016

Structural model of amyloid fibrils for amyloidogenic peptide from Bgl2p-glucantransferase of S. cerevisiae cell wall and its modifying analog. New morphology of amyloid fibrils.

Biochim Biophys Acta 2016 11 6;1864(11):1489-99. Epub 2016 Aug 6.

Institute of Protein Research, Russian Academy of Science, 142290 Pushchino, Moscow Region, Russia. Electronic address:

We performed a comparative study of the process of amyloid formation by short homologous peptides with a substitution of aspartate for glutamate in position 2 - VDSWNVLVAG (AspNB) and VESWNVLVAG (GluNB) - with unblocked termini. Peptide AspNB (residues 166-175) corresponded to the predicted amyloidogenic region of the protein glucantransferase Bgl2 from the Saccharomyces cerevisiae cell wall. The process of amyloid formation was monitored by fluorescence spectroscopy (FS), electron microscopy (EM), tandem mass spectrometry (TMS), and X-ray diffraction (XD) methods. The experimental study at pH3.0 revealed formation of amyloid fibrils with similar morphology for both peptides. Moreover, we found that the morphology of fibrils made of untreated ammonia peptide is not mentioned in the literature. This morphology resembles snakes lying side by side in the form of a wave without intertwining. Irrespective of the way of the peptide preparation, the rate of fibril formation is higher for AspNB than for GluNB. However, preliminary treatment with ammonia highly affected fibril morphology especially for AspNB. Such treatment allowed us to obtain a lag period during the process of amyloid formation. It showed that the process was nucleation-dependent. With or without treatment, amyloid fibrils consisted of ring-like oligomers with the diameter of about 6nm packed either directly ring-to-ring or ring-on-ring with a slight shift. We also proposed the molecular structure of amyloid fibrils for two studied peptides.
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http://dx.doi.org/10.1016/j.bbapap.2016.08.002DOI Listing
November 2016

X-ray diffraction and electron microscopy data for amyloid formation of Aβ40 and Aβ42.

Data Brief 2016 Sep 20;8:108-13. Epub 2016 May 20.

Institute of Protein Research, Russian Academy of Sciences, 142290 Pushchino, Moscow Region, Russia.

The data presented in this article are related to the research article entitled "One of the possible mechanisms of amyloid fibrils formation based on the sizes of primary and secondary folding nuclei of Aβ40 and Aβ42" (Dovidchenko et al., 2016) [1]. Aβ peptide is one of the most intensively studied amyloidogenic peptides. Despite the huge number of articles devoted to studying different fragments of Aβ peptide there are only several papers with correct kinetics data, also there are a few papers with X-ray data, especially for Aβ42. Our data present X-ray diffraction patterns both for Aβ40 and Aβ42 as well for Tris-HCl and wax. Moreover, our data provide kinetics of amyloid formation by recombinant Аβ40 and synthetic Аβ42 peptides by using electron microscopy.
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http://dx.doi.org/10.1016/j.dib.2016.05.020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4889875PMC
September 2016

Insulin and Lispro Insulin: What is Common and Different in their Behavior?

Curr Protein Pept Sci 2017 ;18(1):57-64

Institute of Protein Research, Russian Academy of Sciences, 142290 Pushchino, Moscow Region, Russia.

There are different insulin analogues with various pharmacokinetic characteristics, such as, rapid-acting, long-acting, or intermediate-acting analogues. Since insulin tends to form amyloid aggregates, it is of particular interest to measure characteristic times of formation of amyloid aggregates and compare those to action times for insulin and its analogues. For the study we have chosen one of the insulin analogues - insulin Lispro, which is a fast acting insulin analog. It is usually thought of amyloid aggregation as a nucleation-dependent process. We have estimated the size of the primary nucleus to be one monomer and the size of the secondary nucleus to be around zero in both insulin and Lispro insulin aggregation processes. The main structural element of insulin and Lispro insulin amyloid fibrils is a rounded ring oligomer of about 6-7 nm in diameter, about 2-3 nm in height and about 2 nm in diameter of the hole. Fibrils of several μm in length are produced due to interaction of such oligomers. The packing of ring oligomers in fibrils differs because of the difference in their orderliness. Though the initial stages of fibril formation (monomer, oligomer) are similar, the further process depends on the unique sequence of each peptide. Namely the sequence affects the final morphology of mature amyloids. These observations allow us to conclude that formation of fibrils by short peptides occurs via and by means of oligomer ring structures. Such an important issue as the nature of polymorphism of insulin amyloid fibrils has been settled by us. The role of early oligomeric aggregates in such processes as nucleation and aggregation of amyloid fibrils has been examined.
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http://dx.doi.org/10.2174/1389203717666160526122421DOI Listing
February 2017

Smooth muscle titin forms in vitro amyloid aggregates.

Biosci Rep 2016 07 20;36(3). Epub 2016 May 20.

Laboratory of Structure and Functions of Muscle Proteins, Institute of Theoretical and Experimental Biophysics, Russian Academy of Sciences, 142290 Pushchino, Moscow Region, Russian Federation.

Amyloids are insoluble fibrous protein aggregates, and their accumulation is associated with amyloidosis and many neurodegenerative diseases, including Alzheimer's disease. In the present study, we report that smooth muscle titin (SMT; 500 kDa) from chicken gizzard forms amyloid aggregates in vitro This conclusion is supported by EM data, fluorescence analysis using thioflavin T (ThT), Congo red (CR) spectroscopy and X-ray diffraction. Our dynamic light scattering (DLS) data show that titin forms in vitro amyloid aggregates with a hydrodynamic radius (Rh) of approximately 700-4500 nm. The initial titin aggregates with Rh approximately 700 nm were observed beyond first 20 min its aggregation that shows a high rate of amyloid formation by this protein. We also showed using confocal microscopy the cytotoxic effect of SMT amyloid aggregates on smooth muscle cells from bovine aorta. This effect involves the disorganization of the actin cytoskeleton and result is cell damage. Cumulatively, our results indicate that titin may be involved in generation of amyloidosis in smooth muscles.
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http://dx.doi.org/10.1042/BSR20160066DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5293577PMC
July 2016

One of the possible mechanisms of amyloid fibrils formation based on the sizes of primary and secondary folding nuclei of Aβ40 and Aβ42.

J Struct Biol 2016 06 22;194(3):404-14. Epub 2016 Mar 22.

Institute of Protein Research, Russian Academy of Sciences, 142290 Pushchino, Moscow Region, Russia. Electronic address:

In the presented paper, theoretical as well as electron microscopy and X-ray diffraction experimental approaches were employed for studding the process of Aβ amyloid formation. Using quantitative estimates of a number of monomers which form the nuclei of amyloid fibrils the sizes of folding nuclei of amyloid fibrils for Aβ40 and 42 have been determined for the first time. We have shown that the size of the primary nucleus of Aβ42 peptide fibrils corresponds to 3 monomers, the size of the secondary nucleus for this peptide is 2 monomers. Applying the same analysis to Aβ40 we conclude that the size of the primary nucleus is 2 monomers, and the size of the secondary nucleus is one monomer. Summation of our theoretical and experimental results has allowed us to propose a new model of the structural organization of amyloid fibrils. Our model suggests that the generation of fibrils takes place along the following simplified pathway: a monomer→a ring oligomer→a mature fibril consisting of ring oligomers. These data shed more light upon our understanding of what sizes of the oligomers could represent main targets for future therapies (tetramers for Aβ42 and trimers for Aβ40), and aid in the development of inhibitors of Aβ40 and 42 oligomer formation.
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http://dx.doi.org/10.1016/j.jsb.2016.03.020DOI Listing
June 2016

Search for conserved amino acid residues of the α-crystallin proteins of vertebrates.

J Bioinform Comput Biol 2016 04 28;14(2):1641004. Epub 2016 Jan 28.

1 Institute of Protein Research, Russian Academy of Sciences, Institutskaya str., 4 Pushchino, Moscow Region 142290, Russia.

[Formula: see text]-crystallin is the major eye lens protein and a member of the small heat-shock protein (sHsp) family. [Formula: see text]-crystallins have been shown to support lens clarity by preventing the aggregation of lens proteins. We performed the bioinformatics analysis of [Formula: see text]-crystallin sequences from vertebrates to find conserved amino acid residues as the three-dimensional (3D) structure of [Formula: see text]-crystallin is not identified yet. We are the first who demonstrated that the N-terminal region is conservative along with the central domain for vertebrate organisms. We have found that there is correlation between the conserved and structured regions. Moreover, amyloidogenic regions also correspond to the structured regions. We analyzed the amino acid composition of [Formula: see text]-crystallin A and B chains. Analyzing the occurrence of each individual amino acid residue, we have found that such amino acid residues as leucine, serine, lysine, proline, phenylalanine, histidine, isoleucine, glutamic acid, and valine change their content simultaneously in A and B chains in different classes of vertebrates. Aromatic amino acids occur more often in [Formula: see text]-crystallins from vertebrates than on the average in proteins among 17 animal proteomes. We obtained that the identity between A and B chains in the mammalian group is 0.35, which is lower than the published 0.60.
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http://dx.doi.org/10.1142/S0219720016410043DOI Listing
April 2016

Studies of Polymorphism of Amyloid-β42 Peptide from Different Suppliers.

J Alzheimers Dis 2015 ;47(3):583-93

Institute of Protein Research, Russian Academy of Science, Pushchino, Moscow Region, Russia.

The aim of this study was to investigate the process of amyloidogenesis of amyloid-β (Aβ)42 peptide, by means of fluorescence spectroscopy, electron microscopy, X-ray diffraction, and mass spectrometry. It has been repeatedly reported in the literature that the process of fibril formation by Aβ42 peptide depends considerably not only upon the specific conditions (ionic conditions, pH, temperature, mixing, etc.), as well as the manufacturing route (synthetic or recombinant), but also on the methods of synthesis and purification. We have, for the first time, systematically analyzed samples of Aβ42 peptide supplied by five different companies (Anaspec, Invitrogen, Enzo, Sigma-Aldrich, and SynthAssist) and obtained evidence of significant variability, including lot to lot variations. All studied samples formed amyloid-like fibrils at pH3-6, and the fibrils contained cross-β structures. Samples from Anaspec, Invitrogen, and Enzo formed one particular type of amyloid-like fibrils, while the samples from Sigma-Aldrich and SynthAssist formed another distinct type of fibrils. The observed polymorphism emphasizes the capacity of the Aβ42 peptide to act as a prion agent with varying structural characteristics. The presented data have allowed us to propose a possible mechanism of formation of amyloid-like fibrils.
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http://dx.doi.org/10.3233/JAD-150147DOI Listing
July 2016

How to determine the size of folding nuclei of protofibrils from the concentration dependence of the rate and lag-time of aggregation. II. Experimental application for insulin and LysPro insulin: aggregation morphology, kinetics, and sizes of nuclei.

J Phys Chem B 2014 Feb 24;118(5):1198-206. Epub 2014 Jan 24.

Institute of Protein Research , Russian Academy of Sciences, 4 Institutskaya str., Pushchino, Moscow Region, 142290, Russia.

Insulin is a commonly used protein for studies of amyloidogenesis. There are a few insulin analogues with different pharmacokinetic characteristics, in particular the onset and duration of action. One of them is LysPro insulin. The behavior of LysPro insulin in the process of amyloid formation has not been studied in detail yet. To quantitatively investigate the differences between insulin and LysPro insulin in the aggregation reaction, we used thioflavin T fluorescence assay, electron microscopy, X-ray diffraction methods, and theoretical modeling. Kinetic experimental data for both insulin samples demonstrated the increase of the lag-time for LysPro insulin at low concentrations of monomers, particularly at 2 and 4 mg/mL, which corresponds to the pharmaceutical concentration. However, the morphology of insulin and LysPro insulin fibrils and their X-ray diffraction patterns is identical. Mature fibrils reach 10-12 μm in length and about 3-4 nm in diameter. The obtained analytical solution allow us to determine the sizes of the primary and secondary nuclei from the experimentally obtained concentration dependences of the time of growth and the ratio of the lag-time duration to the time of growth of amyloid protofibrils. In the case of insulin and LysPro insulin, we have exponential growth of amyloid protofibrils following the "bifurcation + lateral growth" scenario. In accord with the developed theory and the experimental data, we obtained that the size of the primary nucleus is equal to one monomer and the size of the secondary nucleus is zero in both insulin and LysPro insulin.
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http://dx.doi.org/10.1021/jp4083568DOI Listing
February 2014

Formation of amyloid-like fibrils by Y-box binding protein 1 (YB-1) is mediated by its cold shock domain and modulated by disordered terminal domains.

PLoS One 2012 8;7(5):e36969. Epub 2012 May 8.

Group of Protein Biosynthesis Regulation, Institute of Protein Research, Russian Academy of Sciences, Pushchino, Russia.

YB-1, a multifunctional DNA- and RNA-binding nucleocytoplasmic protein, is involved in the majority of DNA- and mRNA-dependent events in the cell. It consists of three structurally different domains: its central cold shock domain has the structure of a β-barrel, while the flanking domains are predicted to be intrinsically disordered. Recently, we showed that YB-1 is capable of forming elongated fibrils under high ionic strength conditions. Here we report that it is the cold shock domain that is responsible for formation of YB-1 fibrils, while the terminal domains differentially modulate this process depending on salt conditions. We demonstrate that YB-1 fibrils have amyloid-like features, including affinity for specific dyes and a typical X-ray diffraction pattern, and that in contrast to most of amyloids, they disassemble under nearly physiological conditions.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0036969PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3348147PMC
September 2012

Compact globular structure of Thermus thermophilus ribosomal protein S1 in solution: sedimentation and calorimetric study.

J Biol Chem 2003 Sep 14;278(38):36311-4. Epub 2003 Jul 14.

Institute of Protein Research, Russian Academy of Sciences, Pushchino, Moscow Region 142290, Russia.

Ribosomal protein S1 of Thermus thermophilus overexpressed in Escherichia coli cells has been isolated and subjected to studies by analytical sedimentation and differential scanning microcalorimetry techniques. It has been demonstrated that the protein of 60 kDa sediments at s020,w = 4.6 S and has the diffusion coefficient D020,w = 6.7 x 10(-7) cm2/s in 25 mm HEPES-NaOH buffer, pH 7.5 (similarly to bovine serum albumin of 66 kDa that sediments at s0 20,w = 4.4 S and D020,w =6.0 x 10(-7) cm2/s), indicating its compact globular conformation under these conditions. The microcalorimetry study has shown the presence of a cooperative tertiary structure melting at 90 degrees C, but with several (probably three) independent cooperative domains. In the presence of 100 mm NaCl the protein becomes more asymmetric (s020,w = 3.1 S) but does not lose its cooperativity and thermostability, this suggesting just the weakening of interdomain ionic interactions. The compact globular conformation of protein S1 seems to be most likely within the ribosome.
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http://dx.doi.org/10.1074/jbc.M304713200DOI Listing
September 2003

Ribosomal protein S1 from Thermus thermophilus: its detection, identification and overproduction.

FEBS Lett 2002 Aug;525(1-3):88

Institute of Protein Research, Russian Academy of Sciences, 142290, Moscow Region, Pushchino, Russia.

Ribosomal protein S1 has been identified in Thermus thermophilus ribosomes. The gene of ribosomal protein S1 from Thermus thermophilus has been cloned and overexpressed in Escherichia coli. A procedure for purification of the protein has been developed.
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http://dx.doi.org/10.1016/s0014-5793(02)03092-2DOI Listing
August 2002