Mapping of antibody binding epitopes of a recombinant Poa p IX allergen

Zhang, Lei; Olsen, Egil; Kisil, F.T.; Hill, Robert D.; Sehon, A.H.; Mohapatra, Shyam S.

doi:10.1016/0161-5890(92)90175-w

Cited by 29 publications

(17 citation statements)

References 24 publications

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“…Furthermore, the antibody binding sites of the Poa p 5 recombinant allergens, rKBG60, and the con served C-terminal fragment encoded by cDNA. KBG8.3 [16], were defined using overlapping cDNA clones [15], peptides synthesized on polypropylin pins [17] and synthet ic peptides with the aid of murine antisera to 1KBG6O [18], Moreover, our previous studies with the recombinant [3-galactosidase-KBG8.3 chimeric protein indicated that adju vants played a significant role in modulating the develop ment of specific immune response to the recombinant KBG allergen [19], In an effort to gain further insight into the ef fect of host genetic and adjuvant factors in shaping the im mune response to the recombinant Poa p 5 allergen, in this study we have examined fine specificity o f antibodies in duced by rKBG8.3 in six different strains and with two dif ferent adjuvants utilizing overlapping peptides encompass ing this allergen. Our results demonstrate that host genetic background and adjuvant factors significantly influence not only the isotype specificity but also fine specificity of anti bodies elicited to the recombinant KBG allergen.…”

Section: Introductionmentioning

confidence: 99%

Host Genetic and Adjuvant Factors Influence Epitope Specificity to a Major Recombinant Grass Allergen

Yang

Wang²,

Zhang³

et al. 1996

Int Arch Allergy Immunol

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The role of host genetic and adjuvant factors in the induction of immune responses to a major recombinant Kentucky bluegrass allergen was examined utilizing five strains of mice and two different adjuvants. Analysis of the recombinant allergen-specific antibodies induced in these strains indicated that the antibodies of various isotypes were differentially regulated. In terms of IgE antibody response, BDF1 and DBA/2 were characterized as high responder, whereas BALB/C, CBA/J and C57BL/6 were intermediate and SJL was a low responder. In different strains, both dextran sulfate (DS) and complete Freund’s adjuvant (CFA), as adjuvants, induced recombinant allergen-specific IgE antibodies of similar titer, however, CFA induced higher IgG2a and lower IgM antibodies compared to DS. Further, analysis of T cell proliferative responses of the splenocytes of different strains demonstrated that these strains varied also in their capacity to respond to synthetic peptides. Furthermore, utilizing a panel of synthetic peptides corresponding to the recombinant allergen, we demonstrated that the antibodies induced by the recombinant allergen with CFA in different strains vary with respect to their epitope specificity. In the BDF1 strain, compared to DS, CFA as adjuvant induced recombinant allergen-specific antibodies of additional peptide specificity. Taken together, these results suggest that both host genetic background and adjuvants govern the fine specificity of antibodies produced against this recombinant Kentucky bluegrass allergen.

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Section: Introductionmentioning

confidence: 99%

Host Genetic and Adjuvant Factors Influence Epitope Specificity to a Major Recombinant Grass Allergen

Yang

Wang²,

Zhang³

et al. 1996

Int Arch Allergy Immunol

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“…However, it is difficult to interpret such results in the context of the allergy because of the variations in affinity of diverse antibodies, and the presence of antiiditypic and specific IgG antibodies in vivo. As our previous study using 10-residue peptides had suggested that some of the epitopes on the rKBG60 may be sequential, 25 we examined the immunoglobulin-binding property of a complete set of 20-residue, 10-overlapping synthetic peptides representing rKBG60. We elected to take a polyclonal approach and employed an ELISAbased assay to examine the antibody binding ability of these peptides.…”

Section: Discussionmentioning

confidence: 99%

“…Furthermore, the antibodies induced by peptides 25-28 were cross-reactive with each other, which is consistent with our previous finding that this hydrophobic part of the molecule is highly immunogenic. 25 To elucidate peptides recognized by human T cells, we used a PBMC proliferation assay rather than T-cell lines or clones. The results demonstrated that rKBG60 allergen contained a number of T-cell epitopes that varied with each individual.…”

Section: Multiple Epitopes On a Major Grass Allergen 287mentioning

confidence: 99%

“…However, we failed to detect human IgE-binding epitopes using the decapeptides on the pins. 25 In the present study, we have extended our analysis of B-and T-cell epitopes utilizing 20-residue long synthetic peptides, each with a 10-residue overlap, for further defining these epitopes in humans and mice.…”

Section: Introductionmentioning

confidence: 99%

“…We have previously reported molecular cloning and sequencing of a grass allergen, rKBG60, from Kentucky Bluegrass (Poa pratensis) pollen. [23][24][25][26] Furthermore, the antibody-binding sites of rKBG60 have been defined using overlapping cDNA clones and peptides synthesized on polypropylin pins with the aid of murine antisera to rKBG60. However, we failed to detect human IgE-binding epitopes using the decapeptides on the pins.…”

Section: Introductionmentioning

confidence: 99%

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Multiple B‐ and T‐cell epitopes on a major allergen of Kentucky Bluegrass pollen

Zhang¹,

Yang

Chong³

et al. 1996

Immunology

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SUMMARYThe B-and T-cell epitopes of a recombinant grass allergen, rKBG60, were delineated using a set of overlapping synthetic peptides. Direct binding by enzyme-linked immunosorbent assay (ELISA) utilizing serum pools led to the identification of 13 murine immunoglobulin-, and nine to 13 human IgGand five to seven human IgE-reactive overlapping peptides. Of the peptides which bound to human IgE antibodies, all but three peptides bound to human and/or murine IgG antibodies. Furthermore, eight out of 12 synthetic peptides induced antigen-specific antibodies in mice, suggesting that these peptides contained epitopes that recognized and/or induced T cells. These results, in conjunction with crossrecognition of different peptides at the C-terminus of rKBG60 by antibodies to neighbouring or nonoverlapping peptides suggest that the C-terminus of this antigen represents a dominant antigenic and allergenic site. Peripheral blood mononuclear cell (PBMC) proliferation studies using these synthetic peptides for 13 grass allergic individuals indicated that seven potential human T-cell epitopes exist on this allergen. Taken together, the results demonstrate that multiple B-and T-cell epitopes exist on this major group of grass allergens, the majority of which are localized at the C-terminus of this antigen.

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Comparison of the antibody response to bee venom phospholipase A2 induced by natural exposure in humans or by immunization in mice

Schneider¹,

Dudler²,

Annand

et al. 1997

J. Mol. Recognit.

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Two human and twelve murine monoclonal antibodies directed against the main bee venom allergen phospholipase A2 (PLA) were evaluated for their fine specificity of binding to antigen and their ability to inhibit the enzymatic activity of the antigen. Antibodies were induced by natural exposure of beekeepers to bee venom or immunization of mice via different methods. Both human monoclonal antibodies (hmAbs) were previously shown to recognize the native three-dimensional conformation of PLA and are directed against discontinuous epitopes which include lysine residue at position 25 as a contact residue. In contrast, six of the murine monoclonal antibodies (mmAbs) bind to the denatured structure of the protein as determined by enzyme-linked immunosorbent assay. The epitopes recognized are located near the C-terminal end (n = 8), in the centre of the polypeptide (n = 1), near the N-terminal end (n = 1) or include the carbohydrate part (n = 2) of the PLA molecule. The capacity of the antibodies to modify the enzymatic activity was also determined. The hmAbs significantly inhibit the enzyme (70-79%), whereas the mmAbs produced various degrees of inhibition (39-100%). Since the X-ray structure of PLA is known, the epitopes can be visualized in the context of the three-dimensional structure of the antigen. A qualitative correlation was found between the location of epitopes and the inhibition pattern. Strong inhibition was seen with those antibodies that recognize epitopes that lie on the surface of the enzyme that is thought to contact the phospholipid bilayer. The results show that even though both hmAbs and most mmAbs inhibit the enzymatic activity of PLA, the antigen-binding properties of antibodies from different species raised after different routes of immunization differ significantly. Thus, detailed epitope mapping studies using murine antibodies prepared by artificial immunization may have limited value in predicting epitope patterns relevant to an antibody response to allergens in humans naturally exposed to antigen/allergen.

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Mapping of antibody binding epitopes of a recombinant Poa p IX allergen

Cited by 29 publications

References 24 publications

Host Genetic and Adjuvant Factors Influence Epitope Specificity to a Major Recombinant Grass Allergen

Host Genetic and Adjuvant Factors Influence Epitope Specificity to a Major Recombinant Grass Allergen

Multiple B‐ and T‐cell epitopes on a major allergen of Kentucky Bluegrass pollen

Comparison of the antibody response to bee venom phospholipase A2 induced by natural exposure in humans or by immunization in mice

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