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11.
Bogachek MV Protopopova EV Ternovoĭ VA Kachko AV Ivanova AV Ivanisenko VA Loktev VB 《Molekuliarnaia biologiia》2005,39(5):813-822
Fragments cDNA (nt 935-1475, 1091-1310, 935-1193) encoding N-terminal part of protein E of West Nile virus (WNV), strain LEIV-Vlg99-27889-human were obtained and cloned. Recombinant polypeptides of glycoprotein E (E1-86, E53-126, E1-180) of the WNV with corresponding amino acid sequence to the cloned fragments of cDNA and modeling the epitopes of domains I and II of surface glycoprotein E were purified by affinity chromatography. Twelve types of monoclonal antibodies (MAbs) created in our laboratory against recombinant polypeptide E1-180 interact with glycoprotein E of the WNV as results of Western blot and ELISA that is demonstrating an similarity of chemical structure of short recombinant polypeptides and corresponding amino acid sequence regions of WNV protein E. Analysis of interactions of MAbs with short recombinant polypeptides and protein E of tick-borne encephalitis virus let us reveal no less than six epitopes within domains I and II of glycoprotein E of the WNV. No less than seven types of MAbs to 86-126 aa region of the domain II were found where located peptide providing fusion of virus--cell membranes (98-110 aa). The epitope for anti-receptor MAbs 10H10 within 53-86 aa region of domain II of protein E of the WNV was mapped and it shows that the fusion peptide and co-receptor of protein E for cellular laminin-binding protein (LBP) are spatial nearness. X-ray model of protein E let us suppose that bc-loop (73-89 aa) of domain II interacts with LBP and together with cd-loop (fusion peptide) determines an initial stages of penetration virions into cell. 相似文献
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B. G. Gavrilov G. S. Monastyrskaya T. V. Velikodvorskaya O. B. Filyukova S. N. Konovalova A. A. Kachko E. V. Protopopova L. G. Nikolaev V. B. Loktev E. D. Sverdlov 《Russian Journal of Bioorganic Chemistry》2003,29(2):154-159
The genes that were induced and suppressed in human embryonic kidney cell line RH upon the infection with tick-borne encephalitis virus were studied by the method of subtractive hybridization. The expression of interferon-induced genes IFI-54K and IFI-56K in the infected cells was found to increase 50–100-fold. 相似文献
13.
M. V. Bogachek E. V. Protopopova V. A. Ternovoi A. V. Kachko A. V. Ivanova V. A. Ivanisenko V. B. Loktev 《Molecular Biology》2005,39(5):710-718
Complementary DNA fragments (nucleotides 935–1475, 1091–1310, and 935–1193) encoding the N-terminal portion of glycoprotein E of West Nile virus (WNV), strain LEIV-Vlg99-27889-human, were cloned. Recombinant polypeptides of glycoprotein E (E1–180, E53–126, and E1–86) of the WNV having amino acid sequences corresponding to the cloned cDNA fragments and mimicking the main functional regions of domains I and II of surface glycoprotein E were purified by affinity chromatography. According to ELISA and Western blotting, 12 types of monoclonal antibodies (MAbs) raised in our laboratory against recombinant polypeptide E1–180 recognized the WNV glycoprotein E. This is indicative of similarity between the antigenic structures of the short recombinant polypeptides and corresponding regions of the glycoprotein. Analysis of interactions of the MAbs with short recombinant polypeptides and protein E of tick-borne encephalitis virus revealed at least six epitopes within domains I and II of the WNV protein E. We found at least seven MAb types against the region between amino acid residues (aa) 86 and 126 of domain II, which contains the peptide responsible for fusion of the virus and cell membranes (residues 98–110). The epitope for antireceptor MAbs 10H10 was mapped within the 53–86 aa region of domain II of WNV protein E, which is evidence for the spatial proximity of the fusion peptide and the coreceptor of protein E (residues 53–86) for cellular laminin-binding protein (LBP). The X-ray pattern of protein E suggests that the bc loop (residues 73–89) of domain II interacts with LBP and, together with the cd loop (fusion peptide), determines the initial stages of flavivirus penetration into the cell. 相似文献
14.
Gavrilov BG Monastyrskaia GS Velikodvorskaia TV Filiukova OB Konovalova SN Kachko AA Protopopova EV Nikolaev LG Loktev VB Sverdlov ED 《Bioorganicheskaia khimiia》2003,29(2):175-180
The genes that were induced and suppressed in human embryonic kidney cell line RH upon the infection with tick-borne encephalitis virus were studied by the method of subtractive hybridization. The expression of interferon-induced genes IFI-54K and IFI-56K in the infected cells was found to increase 50-100-fold. 相似文献
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M. V. Bogachek B. N. Zaitsev S. K. Sekatskii E. V. Protopopova V. A. Ternovoi A. V. Ivanova A. V. Kachko V. A. Ivanisenko G. Dietler V. B. Loktev 《Biochemistry. Biokhimii?a》2010,75(4):472-480
Recombinant polypeptide containing the 260–466 amino acid sequence of West Nile virus (WNV) strain LEIV-Vlg99-27889-human
glycoprotein E (gpE, E260–466) was constructed. Immunochemical similarity between the E260–466 and gpE of WNV was proven by enzyme immunoassay (EIA), immunoblot, competitive EIA, hemagglutination inhibition, and neutralization
tests using polyclonal and monoclonal antibodies against the viral gpE and recombinant E260–466. Polypeptide E260–466 induced formation of virus neutralizing and cross-reactive antibodies that were interactive with various epitopes of this
recombinant protein. It is shown by evaluation of the interaction of E260–466 with one of the proposed cell receptors of WNV that average E260–466-αVβ3 integrin-specific interaction force measured using atomic force spectroscopy was 80 and 140 pN for single and double
interactions, correspondingly. Taken together with previously described interaction between laminin-binding protein (LBP)
and WNV gpE domain II, it is proposed that WNV gpE can interact specifically with two cellular proteins (LBP and αVβ3 integrin)
during virus entry. 相似文献
17.
Bondarenko EI Protopopova EV Konovalova SN Sorokin AV Kachko AV Surovtsev IV Loktev VB 《Molekuliarnaia genetika, mikrobiologiia i virusologiia》2003,(4):36-39
The interaction of the VEE virus virions with human LBP was investigated. The affinely purified 43 kDa recombinant LBP (rLBP) of man was found to interact effectively with the VEE virus virions purified in immune enzyme assay. The affinity constant of 43 kDa rLBP with virions was equal to 4.3-4.8 x 10(7) M-1. The rabbit antiviral polyclonal antibodies blocked the interaction of rLBP with the VEE virus virions. According to Western blot, rLBP is capable of interacting with the E1 glycoprotein of the VEE virus, which suggests the presence of a specific epitope of binding with LBP in the surface of the E1-E2 heterodimer of the VEE virus. The results confirm that human LBP could be a receptor for the VEE virus. 相似文献
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A V Kachko T B Cheusova A V Sorokin E I Kazachinskaia I O Cheshenko E F Belanov A A Bukreev A V Ivanova I A Razumov E I Riabchikova S V Netesov 《Molekuliarnaia biologiia》2001,35(3):492-499
The full-length gene for Marburg virus (MV) nucleoprotein (NP) was cloned in prokaryotic pQE32 under the control of the T5 promoter and in eukaryotic pTM1 under the control of the promoter for T7 RNA polymerase. Recombinant NP was synthesized in Escherichia coli and in human kidney cell line 293 cotransfected with recombinant vaccinia virus vTF7-3 expressing T7 RNA polymerase. On evidence of electron microscopy with immune detection, recombinant NP formed tubules of two types in E. coli and of a single type in cell line 293. ELISA and immunoblotting with polyclonal and monoclonal antibodies revealed common antigenic determinants in recombinant NP and natural MV NP. 相似文献
20.
Gridina MM Protopopova EV Kachko AV Ivanova AV Bondarenko EI Loktev VB 《Tsitologiia》2007,49(11):966-972
The level of laminin-binding protein (LBP) expression on cellular membranes was studied in three cell lines including 293 cells transformed by plasmide with human LBP gene. Vero cells show a high level of LBP on the cell surfaces and demonstrate a high level of the Venezuelan equine encephalomyelitis (VEE) virus replication. The inhibition of VEE virus replication was more than 200 times as much after treatment of Vero cell surfaces with monoclonal antibodies to human LBP. 293 cells have more low level of LBP on their surfaces but being transformed by plasmide with LBP human gene these cells showed an increase in the level of cellular LBP. The VEE virus replication in transformed cells (9S2) was more than 2000 times higher compared to 293 cells. The results obtained demonstrate a principal role of cellular LBP in VEE virus entry into mammalian cells. It can be proposed that LBP is a key cellular protein at the early stage of VEE virus replication in cells. So, LBP might be a target protein for development of some new generation of antiviral drugs that would be able to inhibit (enhance) the alphavirus replication in human cells. 相似文献