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035 a| (Sirsi) o648767429;

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040 a| EREc| EREd| UtOrBLW;

049 a| NEHH;

100 1 a| Fitch, Christina D.?| UNAUTHORIZED;

245 14 a| The serine protease from the venom of polistes dominulus contains allergenic epitopes /c| by Christina D. Fitch.;

260 a| [Greenville, N.C.] :b| East Carolina University,c| 2010.;

300 a| 189 pages :b| illustrations (some color), digital, PDF file.;

336 a| text2| rdacontent;

337 a| computer2| rdamedia;

338 a| online resource2| rdacarrier;

490 1 a| ECU Brody School of Medicine thesis;

500 a| Presented to the faculty of the Department of the Interdisciplinary Doctoral Program in Biological Sciences.;

500 a| Advisor: Donald R. Hoffman.;

500 a| Title from PDF t.p. (viewed November 9, 2010).;

502 b| Ph.D.c| East Carolina Universityd| 2010.;

504 a| Includes bibliographical references.;

520 3 a| The protection introduced by immunotherapy to patients with allergies to paper wasps may be partial depending on the sensitizing species. Partial cross-reactivity between major paper wasp groups may be the reason for partial protection. Understanding patient response to venom allergens from the paper wasp groups is important to improve and develop effective treatments. Research has shown that the allergenic serine protease may be important in this partial cross-reactivity. In this study we investigated the importance of epitopes on the allergenic serine protease (Pol d 4) from Polistes dominulus. Recombinant Pol d 4 was expressed in prokaryotic and eukaryotic expression systems. The expression products from the eukaryotic system yielded a protein of larger size than predicted size. Two constructs of Pol d 4 were produced in the prokaryotic expression system and were of expected size. Re-folding of the prokaryotic expressed recombinant proteins was not successful. The major amino acid epitopes on Pol d 4 were predicted. Three mutant constructs were chosen, each containing two amino acid substitutions within a predicted epitopes. IgE binding was assayed by immunoblot assays using sera from European and North American patients. The majority of European patient sera had IgE that bound to the recombinant form of Pol d 4. Statistical analysis showed that there was no significant difference in IgE binding to the various proteins. IgE from American patients bound recombinant Pol d 4. Bromelain was used in immunoblot inhibition to assay possible carbohydrate epitopes. The inhibition assay showed evidence of IgE binding. The serine protease from Polistes gallicus was characterized. Identity between the two serine proteases was high. Evidence supports the presence of carbohydrate on the recombinant protease and carbohydrate epitopes may be present. Visually, all three mutant proteins bound IgE; mutant 2 bound IgE from the least number of patients suggesting it may be an important epitope on Pol d 4. Analysis of densitometric data showed no significant difference between the proteins tested. The data suggests that there may be amino acid epitopes present. Further, the high sequence identity between Pol d 4 and Pol g 4 support high cross-reactivity between sister species.;

538 a| System requirements: Adobe Reader.;

538 a| Mode of access: World Wide Web.;

650 2 a| Wasp Venoms.=| ^A1032923;

650 2 a| Epitopesx| toxicity.=| ^A1032772;

653 a| Biology, Molecular;

700 1 a| Hoffman, Donald R.=| ^A1024891;

710 2 a| East Carolina University.b| Department of Interdisciplinary Biological Sciences.?| UNAUTHORIZED;

830 0 a| ECU Brody School of Medicine thesis.=| ^A964744;

856 40 z| Access via ScholarShip u| http://hdl.handle.net/10342/2695;

949 a| Click on web address w| ASIS h| JOYNER101;

949 a| Click on web address w| ASIS h| HSL111;

994 a| C0b| NEH;

096 a| WD 430;

596 a| 1 4;

998 a| 2284734;

999 a| CLICK ON WEB ADDRESSw| ASISc| 1i| 2284734-1001l| JNETm| JOYNERr| Ys| Yt| JNE3ETDu| 7/19/2010x| ETDz| JERESOURCE;

999 a| CLICK ON WEB ADDRESSw| ASISc| 1i| 2284734-2001l| HSLELECm| HSLr| Ys| Yt| HEETDu| 7/19/2010x| ETDz| HERESOURCE;

The serine protease from the venom of polistes dominulus contains allergenic epitopes / by Christina D. Fitch.

Author/creator	Fitch, Christina D.
Other author	Hoffman, Donald R.
Other author	East Carolina University. Department of Interdisciplinary Biological Sciences.
Format	Theses and dissertations
Publication Info	[Greenville, N.C.] : East Carolina University, 2010.
Description	189 pages : illustrations (some color), digital, PDF file.
Supplemental Content	Access via ScholarShip
Subjects	Wasp Venoms. Epitopes--toxicity.

Series	ECU Brody School of Medicine thesis ECU Brody School of Medicine thesis. ^A964744
Summary	The protection introduced by immunotherapy to patients with allergies to paper wasps may be partial depending on the sensitizing species. Partial cross-reactivity between major paper wasp groups may be the reason for partial protection. Understanding patient response to venom allergens from the paper wasp groups is important to improve and develop effective treatments. Research has shown that the allergenic serine protease may be important in this partial cross-reactivity. In this study we investigated the importance of epitopes on the allergenic serine protease (Pol d 4) from Polistes dominulus. Recombinant Pol d 4 was expressed in prokaryotic and eukaryotic expression systems. The expression products from the eukaryotic system yielded a protein of larger size than predicted size. Two constructs of Pol d 4 were produced in the prokaryotic expression system and were of expected size. Re-folding of the prokaryotic expressed recombinant proteins was not successful. The major amino acid epitopes on Pol d 4 were predicted. Three mutant constructs were chosen, each containing two amino acid substitutions within a predicted epitopes. IgE binding was assayed by immunoblot assays using sera from European and North American patients. The majority of European patient sera had IgE that bound to the recombinant form of Pol d 4. Statistical analysis showed that there was no significant difference in IgE binding to the various proteins. IgE from American patients bound recombinant Pol d 4. Bromelain was used in immunoblot inhibition to assay possible carbohydrate epitopes. The inhibition assay showed evidence of IgE binding. The serine protease from Polistes gallicus was characterized. Identity between the two serine proteases was high. Evidence supports the presence of carbohydrate on the recombinant protease and carbohydrate epitopes may be present. Visually, all three mutant proteins bound IgE; mutant 2 bound IgE from the least number of patients suggesting it may be an important epitope on Pol d 4. Analysis of densitometric data showed no significant difference between the proteins tested. The data suggests that there may be amino acid epitopes present. Further, the high sequence identity between Pol d 4 and Pol g 4 support high cross-reactivity between sister species.
General note	Presented to the faculty of the Department of the Interdisciplinary Doctoral Program in Biological Sciences.
General note	Advisor: Donald R. Hoffman.
General note	Title from PDF t.p. (viewed November 9, 2010).
Dissertation note	Ph.D. East Carolina University 2010.
Bibliography note	Includes bibliographical references.
Technical details	System requirements: Adobe Reader.
Technical details	Mode of access: World Wide Web.

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