Search results

1

The construction of the hybrid plasmid containing genes of the central part of phage T4 baseplate and gene 29 product separation

100%

Nieradko J., Szewczyk B.

Acta Biochimica Polonica

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1988

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vol. 35

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issue 4

357-366

2

Introduction to molecular biology of influenza a viruses

81%

Szewczyk B., Bieńkowska-Szewczyk K., Król E.

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2014

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vol. 61

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issue 3

397-401

EN

This minireview presents an overview of current knowledge on virion structure, genome organization and basic events in the development of influenza A virus. The processes of entry, transcription/replication and viral release are described. In this context, the roles of viral proteins (including recently discovered minor polypeptides) in the subsequent stages of viral development are also discussed.

3

Effect of N-glycosylation inhibition on the synthesis and processing of classical swine fever virus glycoproteins

81%

Tyborowska J., Zdunek E., Szewczyk B.

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2007

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vol. 54

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issue 4

813-819

EN

Classical swine fever virus (CSFV) is often used as a surrogate model in molecular studies of the closely related hepatitis C virus. In this report we have examined the effect of the inhibition of glycosylation on the survival and maturation of CSFV. Viral glycoproteins (Erns, E1, E2) form biologically active complexes - homo- and heterodimers, which are indispensable for viral life cycle. Those complexes are highly N-glycosylated. We studied the influence of N-glycosylation on dimer formation using Erns and E2 glycoproteins produced in insect cells after infection with recombinant baculoviruses. The glycoproteins were efficiently synthesized in insect cells, had similar molecular masses and formed dimers like their natural counterparts. Surprisingly, the addition of tunicamycin (an antibiotic which blocks early steps of glycosylation) to insect cell culture blocked not only dimer formation but it also led to an almost complete disappearance of E2 even in monomeric form. Tunicamycin did not exert a similar effect on the synthesis and formation of Erns dimers; the dimers were still formed, which suggests that Erns glycan chains are not necessary for dimer formation. We have also found that very low doses of tunicamycin (much lower than those used for blocking N-glycosylation) drastically reduced CSFV spread in SK6 (swine kidney) cell culture and the virus yield. These facts indicate that N-glycosylation inhibitors structurally similar to tunicamycin may be potential therapeutics for the inhibition of the spread of CSFV and related viruses.

4

Expression of bacteriophage T4 minor baseplate proteins in the bacteriophage T7 promoter/RNA polymerase expression system

81%

Schlichtholtz B., Nieradko J., Gajek M., Szewczyk B.

Acta Biochimica Polonica

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1991

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vol. 38

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issue 3

311-319

5

Antivirals - current trends in fighting influenza

81%

Król E., Rychłowska M., Szewczyk B.

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EN

Influenza virus infection is a major source of morbidity and mortality worldwide. Due to the variable effectiveness of existing vaccines, especially in the early stages of an epidemic, antiviral drugs represent the first line of defense against the virus. Currently, there are two major classes of anti-influenza drugs approved by the FDA for clinical use: M2 protein inhibitors (amantadine and rimantadine) and neuraminidase inhibitors (zanamivir and oseltamivir). However, increasing resistance to these available influenza antivirals among circulating influenza viruses highlights the need to develop alternative approaches for the prevention and/or treatment of influenza. This review presents an overview of currently available drugs for influenza treatment as well as summarizes some new antiviral strategies that are now being tested covering agents targeting both the viral proteins and the host-virus interaction. We discuss their mechanisms of action, resistance and the therapeutic potential as new antiviral drug for use in future influenza pandemics. Additionally, combination therapy based on these drugs is also described.

6

Expression of genes coding for animal virus glycoproteins in heterologous systems

80%

Bieńkowska-Szewczyk K., Szewczyk B.

Acta Biochimica Polonica

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1999

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vol. 46

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issue 2

325-339

7

Recombinant VP60 in the form of virion-like particles as a potential vaccine against rabbit hemorrhagic disease virus*

71%

Gromadzka B., Szewczyk B., Konopa G., Fitzner A., Kęsy A.

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2006

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vol. 53

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issue 2

371-376

EN

Rabbit hemorrhagic disease virus (RHDV) which causes a highly contagious disease of wild and domestic rabbits belongs to the family Caliciviridae. It is a small, positive single-stranded RNA virus with a genome of 7.5 kb and has a diameter of approximately 40 nm. In negatively stained electron micrographs the virus shows typical calicivirus morphology with regularly arranged cup-shaped structures on the surface. It is a major pathogen of rabbits in many countries. Vp60 - a coat protein of molecular mass around 60 kDa is the major antigen of RHDV. It is present as 90 dimeric units per virion particle. We have expressed VP60 gene in the baculovirus system with the aim to use it as a potential vaccine against RHDV and a diagnostic reagent in immunological tests. cDNA of the vp60 gene of strain SGM, was cloned into a baculovirus transfer vector as full-length gene, as well as truncated gene lacking 600 5´-terminal nucleotides. The sequence of SGM VP60 differed markedly from that of the reference strain. Full-length recombinant VP60 protein from the SGM strain self-assembled to form virus-like particles (VLPs). These particles observed by electron microscopy were morphologically similar to native virions and were able to agglutinate human group 0 erythrocytes. After immunization the recombinant particles induced RHDV-specific antibodies in rabbits and guinea pigs. Rabbits immunized with the VLPs were fully protected against challenge with a virulent RHDV.

8

Detection and differentiation of Newcastle disease virus and influenza virus by using duplex real-time PCR

71%

Nidzworski D., Wasilewska E., Smietanka K., Szewczyk B., Minta Z.

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2013

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vol. 60

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issue 3

475-480

EN

Newcastle disease virus (NDV), member of the Paramyxoviridae family and avian influenza virus (AIV), member of the Orthomyxoviridae family, are two main avian pathogens causing serious economic problems in poultry health. Both are enveloped, single-stranded, negative-sense RNA viruses and cause similar symptoms, ranging from sub-clinical infections to severe diseases, including decrease in egg production, acute respiratory syndrome, and high mortality. Similar symptoms hinder the differentiation of infection with the two viruses by standard veterinary procedures like clinical examination or necropsy. To overcome this problem, we have developed a new duplex real-time PCR assay for the detection and differentiation of these two viruses. Eighteen NDV strains, fourteen AIV strains, and twelve other (negative control) strains viruses were isolated from allantoic fluids of specific pathogen-free (SPF), embryonated eggs. Four-weeks-old SPF chickens were co-infected with both viruses (NDV - LaSota and AIV - H7N1). Swabs from cloaca and trachea were collected and examined. The results obtained in this study show that by using duplex real-time PCR, it was possible to detect and distinguish both viruses within less than three hours and with high sensitivity, even in case a bird was co-infected. Additionally, the results show the applicability of the real-time PCR assay in laboratory practice for the identification and differentiation of Newcastle disease and influenza A viruses in birds.

9

Hemagglutinin stalk domain from H5N1 strain as a potentially universal antigen

71%

Uranowska K., Tyborowska J., Jurek A., Szewczyk B., Gromadzka B.

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2014

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vol. 61

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issue 3

541-550

EN

Influenza A virus infections are the major public health concern and cause significant morbidity and mortality each year worldwide. Vaccination is the main strategy of influenza epidemic prevention. However, seasonal vaccines induce strain-specific immunity and must be reformulated annually based on prediction of the strains that will circulate in the next season. Thus, it is essential to develop vaccines that would induce broad and persistent immunity to influenza viruses. Hemagglutinin is the major surface antigen of the influenza virus. Recent studies revealed the importance of HA stalk-specific antibodies in neutralization of different influenza virus strains. Therefore, it is important to design an immunogen that would focus the immune response on the HA stalk domain in order to elicit neutralizing antibodies. In the present study, we report characterization of a conserved truncated protein, potentially a universal influenza virus antigen from the H5N1 Highly Pathogenic Avian Influenza A virus strain. Our results indicate that exposure of the HA stalk domain containing conserved epitopes results in cross reactivity with different antibodies (against group 1 and 2 HAs). Additionally, we conclude that HA stalk domain contains not only conformational epitopes recognized by universal FI6 antibody, but also linear epitopes recognized by other antibodies.

10

Expresion of animal virus genes using Baculovirus AcNPV

61%

Kochan G., Tyborowska J., Szewczyk B.

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vol. 40

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issue 1

1-3

11

Expression of avian influenza haemagglutinin (H5) and chicken interleukin 2 (chIL-2) under control of the ptcB promoter in Lactococcus lactis

52%

Szatraj K., Szczepankowska A., Sączyńska V., Florys K., Gromadzka B., Łepek K., Płucienniczak G., Szewczyk B., Zagórski-Ostoja W., Bardowski J.

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2014

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vol. 61

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issue 3

609-614

EN

Gram-positive and nonpathogenic lactic acid bacteria (LAB) are considered to be promising candidates for the development of new, safe systems of heterologous protein expression. Recombinant LAB has been shown to induce specific local and systemic immune response against selected pathogens, and could be a good alternative to classical attenuated carriers. The main goal of our study was to express the avian influenza haemagglutinin (H5) and chicken interleukin 2 (chIL-2) in Lactococcus lactis. Results of this study were anticipated to lead to construction of lactococcal strain(s) with potential vaccine properties against the avian influenza A (H5N1) virus. Expression of the cloned H5 gene, its His-tagged variant and chIL-2 gene, under the control of the ptcB gene promoter was attested by RT-PCR on transcriptional level and Western or dot blot analysis on translational level, demonstrating that system can be an attractive solution for production of heterologous proteins. The results of the preliminary animal trial conducted in mice are a promising step toward development of a vaccine against avian bird flu using Lactococcus lactis cells as antigen carriers.

12

Characterization of mAb6-9-1 monoclonal antibody against hemagglutinin of avian influenza virus H5N1 and its engineered derivative, single-chain variable fragment antibody

52%

Sawicka R., Siedlecki P., Kalenik B., Radomski J., Sączyńska V., Porębska A., Szewczyk B., Sirko A., Góra-Sochacka A.

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2017

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vol. 64

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issue 1

85-92

EN

Hemagglutinin (HA), as a major surface antigen of influenza virus, is widely used as a target for production of neutralizing antibodies. Monoclonal antibody, mAb6-9-1, directed against HA of highly pathogenic avian influenza virus A/swan/Poland/305-135V08/2006(H5N1) was purified from mouse hybridoma cells culture and characterized. The antigenic specificity of mAb6-9-1 was verified by testing its cross-reactivity with several variants of HA. The mimotopes recognized by mAb6-9-1 were selected from two types of phage display peptide libraries. The comparative structural model of the HA variant used for antibody generation was developed to further facilitate epitope mapping. Based on the sequences of the affinity- selected polypeptides and the structural model of HA the epitope was located to the region near the receptor binding site (RBS). Such localization of the epitope recognized by mAb6-9-1 is in concordance with its moderate hemagglutination inhibiting activity and its antigenic specificity. Additionally, total RNA isolated from the hybridoma cell line secreting mAb6-9-1 was used for obtaining two variants of cDNA encoding recombinant single-chain variable fragment (scFv) antibody. To ensure high production level and solubility in bacterial expression system, the scFv fragments were produced as chimeric proteins in fusion with thioredoxin or displayed on a phage surface after cloning into the phagemid vector. Specificity and affinity of the recombinant soluble and phage-bound scFv were assayed by suitable variants of ELISA test. The observed differences in specificity were discussed.

13

Expression, purification and characterization of glycosylated influenza H5N1 hemagglutinin produced in Pichia pastoris

52%

Kopera E., Dwornyk A., Kosson P., Florys K., Sączyńska V., Dębski J., Cecuda-Adamczewska V., Szewczyk B., Zagórski-Ostoja W., Grzelak K.

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2014

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vol. 61

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issue 3

597-602

EN

The A/swan/Poland/305-135V08/2006 (H5N1-subtype) hemagglutinin (HA) gene was cloned and expressed in yeast Pichia pastoris (P. pastoris). The HA cDNA lacking the C-terminal transmembrane anchor-coding sequence was fused to an α-factor leader peptide and placed under control of the methanol-inducible P. pastoris alcohol oxidase 1 (AOX1) promoter. Two P. pastoris strains: SMD 1168 and KM 71 were used for protein expression. Recombinant HA protein was secreted into the culture medium reaching an approximately 15 mg/L (KM 71 strain). Fusion protein with a His6 tag was purified to homogeneity in one step affinity chromatography. SDS-PAGE and MS/MS analysis indicated that the protein is cleaved into HA1 and HA2 domains linked by a disulfide bond. Analysis of the N-linked glycans revealed that the overexpressed HA is fully glycosylated at the same sites as the native HA in the vaccine strain. Immunological activity of the hemagglutinin protein was tested in mice, where rHA elicited a high immune response.

Refine search results

The construction of the hybrid plasmid containing genes of the central part of phage T4 baseplate and gene 29 product separation

Introduction to molecular biology of influenza a viruses

Effect of N-glycosylation inhibition on the synthesis and processing of classical swine fever virus glycoproteins

Expression of bacteriophage T4 minor baseplate proteins in the bacteriophage T7 promoter/RNA polymerase expression system

Antivirals - current trends in fighting influenza

Expression of genes coding for animal virus glycoproteins in heterologous systems

Recombinant VP60 in the form of virion-like particles as a potential vaccine against rabbit hemorrhagic disease virus*

Detection and differentiation of Newcastle disease virus and influenza virus by using duplex real-time PCR

Hemagglutinin stalk domain from H5N1 strain as a potentially universal antigen

Expresion of animal virus genes using Baculovirus AcNPV

Expression of avian influenza haemagglutinin (H5) and chicken interleukin 2 (chIL-2) under control of the ptcB promoter in Lactococcus lactis

Characterization of mAb6-9-1 monoclonal antibody against hemagglutinin of avian influenza virus H5N1 and its engineered derivative, single-chain variable fragment antibody

Expression, purification and characterization of glycosylated influenza H5N1 hemagglutinin produced in Pichia pastoris