Katerina Zachova

Palacký University of Olomouc, Olmütz, Olomoucký, Czech Republic

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Publications (5)15.62 Total impact

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    ABSTRACT: HIV-1 entry into host cells is mediated by interactions between the virus envelope glycoprotein (gp120/gp41) and host-cell receptors. N-glycans represent approximately 50% of the molecular mass of gp120 and serve as potential antigenic determinants and/or as a shield against immune recognition. We previously reported that N-glycosylation of recombinant gp120 varied, depending on the producer cells, and the glycosylation variability affected gp120 recognition by serum antibodies from persons infected with HIV-1 subtype B. However, the impact of gp120 differential glycosylation on recognition by broadly neutralizing monoclonal antibodies or by polyclonal antibodies of individuals infected with other HIV-1 subtypes is unknown.
    AIDS Research and Therapy 01/2014; 11:23. · 2.54 Impact Factor
  • Michal Krupka, Katerina Zachova, Evzen Weigl, Milan Raska
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    ABSTRACT: Borrelia burgdorferi sensu lato (Spirochaetes) is a group of at least 12 closely related species, some of which are responsible for chronic zoonotic infection that may cause Lyme disease. The only experimentally confirmed vector transmitting Borrelia to mammals is the Ixodes ticks. Borrelia is a highly adapted pathogen that can survive in the host organism in spite of the intense immune responses. Some patients have chronic long-lasting complications despite antibiotic therapy, probably due to adverse effects of the immune responses. A preventive vaccine against this bacterium has not been available due to the relatively broad spectrum and antigenic variability of Borrelia-surface lipoproteins and the different epitope recognition by experimental animals and humans. Although a human vaccine was marketed in the USA, it has been already pulled off the market. In addition, this vaccine was effective only in the USA, where the only pathogenic species is B. burgdorferi sensu stricto. Recent data indicate that a broadly effective vaccine will to be composed of a mixture of several antigens or multiple epitopes.
    Archivum Immunologiae et Therapiae Experimentalis 08/2011; 59(4):261-75. · 2.38 Impact Factor
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    ABSTRACT: Human immunodeficiency virus type 1 (HIV-1) entry is mediated by the interaction between a variably glycosylated envelope glycoprotein (gp120) and host-cell receptors. Approximately half of the molecular mass of gp120 is contributed by N-glycans, which serve as potential epitopes and may shield gp120 from immune recognition. The role of gp120 glycans in the host immune response to HIV-1 has not been comprehensively studied at the molecular level. We developed a new approach to characterize cell-specific gp120 glycosylation, the regulation of glycosylation, and the effect of variable glycosylation on antibody reactivity. A model oligomeric gp120 was expressed in different cell types, including cell lines that represent host-infected cells or cells used to produce gp120 for vaccination purposes. N-Glycosylation of gp120 varied, depending on the cell type used for its expression and the metabolic manipulation during expression. The resultant glycosylation included changes in the ratio of high-mannose to complex N-glycans, terminal decoration, and branching. Differential glycosylation of gp120 affected envelope recognition by polyclonal antibodies from the sera of HIV-1-infected subjects. These results indicate that gp120 glycans contribute to antibody reactivity and should be considered in HIV-1 vaccine design.
    Journal of Biological Chemistry 07/2010; 285(27):20860-20869. · 4.65 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Human immunodeficiency virus type 1 (HIV-1) entry is mediated by the interaction between a variably glycosylated envelope glycoprotein (gp120) and host-cell receptors. Approximately half of the molecular mass of gp120 is contributed by N-glycans, which serve as potential epitopes and may shield gp120 from immune recognition. The role of gp120 glycans in the host immune response to HIV-1 has not been comprehensively studied at the molecular level. We developed a new approach to characterize cell-specific gp120 glycosylation, the regulation of glycosylation, and the effect of variable glycosylation on antibody reactivity. A model oligomeric gp120 was expressed in different cell types, including cell lines that represent host-infected cells or cells used to produce gp120 for vaccination purposes. N-Glycosylation of gp120 varied, depending on the cell type used for its expression and the metabolic manipulation during expression. The resultant glycosylation included changes in the ratio of high-mannose to complex N-glycans, terminal decoration, and branching. Differential glycosylation of gp120 affected envelope recognition by polyclonal antibodies from the sera of HIV-1-infected subjects. These results indicate that gp120 glycans contribute to antibody reactivity and should be considered in HIV-1 vaccine design.
    Journal of Biological Chemistry 05/2010; 285(27):20860-9. · 4.65 Impact Factor
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    ABSTRACT: Heat shock protein 70 kDa (hsp70), a molecular chaperone involved in folding of nascent proteins, has been studied for its ability to activate innate and specific immunity. High purity hsp70 preparation is generally required for immunization experiments, because endotoxins and other immunologically active contaminants may affect immune responses independently of hsp70. We have developed a novel modification of E. coli-expression medium that enabled a simple two-step production and purification method for endotoxin-free recombinant hsp70. During Ni-NTA-based affinity purification of hsp70, a contaminating protein from host E. coli cells, L-glutamine-D-fructose-6-phosphate aminotransferase (GFAT), was identified. By testing various compounds, supplementation of growth medium with a GFAT metabolite, N-acetylglucosamine, was found to reduce GFAT expression and increase the total hsp70 yield five times. The new protocol is based on column purification of His-tagged hsp70 protein produced by E. coli with the modified medium, followed by endotoxin removal by Triton X-114 extraction. This approach yielded hsp70 with high purity and minimal endotoxin contamination, making the final product acceptable for immunization experiments. In summary, a simple modification of growth medium allowed production of recombinant mouse hsp70 in high yield and purity, thus compatible with immunological studies. This protocol may be useful for production of other His.
    Journal of Microbiology and Biotechnology 08/2009; 19(7):727-33. · 1.40 Impact Factor