Report 2008-2010

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Lab Leader Aurelija Žvirblienė, PhD Laboratory of Immunology and Cell Biology phone: 370 5 2602117 e-mail: azvirb@ibt.lt Research Associates Arvydas Kanopka, PhD. Petras Stakėnas, PhD. Milda Plečkaitytė, PhD. Junior Researchers Daiva Bakonytė, M Sc. Rita Lasickienė, M.Sc. Indrė Šėžaitė, M.Sc. Eglė Jakubauskienė, M.Sc. PhD Students Indrė Kučinskaitė-Kodzė, M.Sc. Inga Pečiulienė, M.Sc. Technical Staff Aldona Savičienė Leokadija Diglienė Undergraduate Laurynas Vilys, B.sc Vaida Žilaitytė Sandra Armalytė Dovilė Dekaminavičiūtė, B.sc. Kristina Nalivaiko Ieva Kairytė 35 th anniversary 36
37 The Laboratory of Immunology and Cell Biology consists of three research groups. In 2008-2010, the research was focussed to the following topics: antigenic characterization of recombinant viral and bacterial proteins, development of monoclonal antibodies (Dr. A.Žvirblienė), regulation of gene expression by alternative splicing (Dr. A.Kanopka), molecular epidemiology of tuberculosis (Dr. P.Stakėnas). Antigenicity studies of recombinant viral proteins This work was aimed at evaluating the antigenic properties of yeast-expressed recombinant proteins that might be exploited as potential vaccines or diagnostic tools. Using different immunochemical assays, the antigenic structure of yeast-expressed nucleocapsid (N) proteins of paramyxoviruses, such as human parainfluenza virus type 3 (hPIV3) and Menangle virus has been investigated. Recombinant N proteins were expressed at the Laboratory of Eukaryote Gene Engineering. The yeast-expressed N proteins used in this study were self-assembled A into nucleocapsid-like structures (NLPs) similar to that of native virus. To identify B-cell epitopes, we have employed monoclonal antibodies (MAbs) raised against recombinant N proteins as well as human sera from virus-infected individuals. The localization of B-cell epitopes was studied using recombinant overlapping N protein fragments, PepScan analysis and competitive ELISAs. The majority of MAb epitopes were mapped within the C-termini of N proteins. Cross-inhibition studies with human sera demonstrated similar localization of B cell epitopes recognized by serum antibodies from naturally infected individuals, which revealed a clear antigenic similarity between recombinant and virus-derived N proteins. Our data suggest that different paramyxoviruses may contain highly conserved surface-exposed structures within the C-termini of N proteins that are well accessible to B cells and elicit antibody response. These findings may have important implications for the design of new vaccines and diagnostic reagents (Zvirbliene et al., Viral Immunol. 2009, Zvirbliene et al., Arch Virol. 2010). B Figure 1. Immunochemical detection of virus-infected cells using monoclonal antibodies raised against recombinant N proteins of hPIV3 (A) and Menangle virus (B). Functional and immunological characterization of recombinant vaginolysin Vaginolysin (VLY) is the main virulence factor released by Gardnerella vaginalis. VLY is a protein toxin that acts as a hemolysin. It belongs to the group of cholesterol-dependent cytolysins (CDCs), a large family of related pore-forming toxins found in five different genera of Grampositive bacteria. Generation of MAbs against bacterial cytolysins may promote elucidation of the relationship between their structure and mode of action. In the current study, we have developed and characterized a panel of MAbs against recombinant VLY and demonstrated that several MAbs neutralize VLY in the in vitro hemolytic assay. By using a series of recombinant proteins we have studied the specificity of the MAbs and mapped the epitope recognized by the most potent neutralizing MAb. The epitope for this MAb was localized near the N-terminus of VLY and included the conserved motif (VAARMQYD, aa 189-196) supposed to be involved in VLY oligomerization. Furthermore, we have employed the MAbs in a sandwich ELISA for VLY quantitation. In conclusion, the MAbs described in the current study may be useful for structural and functional studies of VLY as well as immunodetection of VLY in biological specimens (Zvirbliene et al., Toxicon 2010, PCT/LT2009/000005). This work was supported by the Lithuanian State Science and Studies Foundation/Lithuanian Science Council (grant No. N-17) Dr. M.Plečkaitytė and Master student J.Alesiūtė investigate immunochemical properties of vaginolysin Figure 2. Schematic representation of VLY domains (D1-D4). Dashed area indicates VLY region recognized by the neutralizing monoclonal antibody.
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Page 5 and 6: 3 Director’s note This triennial
Page 8 and 9: INSTITUTE OF BIOTECHNOLOGY: The his
Page 10 and 11: IBT citations in 1991 - 2009 ISI We
Page 12 and 13: Grants 1995-2010 FRAMEWORK 7 PROGRA
Page 14 and 15: 1997 1999 2000 2001 2002 2004 Docto
Page 16 and 17: Strengthening and Sustaining the Eu
Page 18 and 19: Lab Leader Virginijus Šikšnys, Ph
Page 20 and 21: Structure and function of restricti
Page 22 and 23: Time-resolved fluorescence studies
Page 24 and 25: Lab Leader Saulius Klimašauskas, P
Page 26 and 27: HPLC-MS analysis performed by Zita
Page 28 and 29: Archaeal C/D box RNPs: towards prog
Page 30 and 31: Lab leader Gintautas Žvirblis, PhD
Page 32 and 33: In conclusion, the dual potential o
Page 34 and 35: For that purpose 93 serum samples w
Page 36 and 37: International Collaboration Prof. D
Page 40 and 41: Master student V.Žilaitytė, Dr. A
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Page 46 and 47: Here at the LBDD we have cloned and
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Page 56 and 57: DNA Sequencing Center Eglė Rudokie
Page 58 and 59: Design by UAB Idea Artis Printed by

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