Upload
maksymilian
View
212
Download
0
Embed Size (px)
Citation preview
J ALLERGY CLIN IMMUNOL
VOLUME 131, NUMBER 2
Abstracts AB19
SATURDAY
67 A Method of Reducing Allergenicity of Legume ProteinsNaveen Arora1, Ramkrashan Kasera2, Anand Singh, PhD3,
Shakuntala Lavasa4, Komerla Nagendra5; 1CSIR- IGIB, Delhi, India,2CSIR-Institute of Genomics and Integrative Biology, India, 3Institute of
Genomics and Integrative Biology, DELHI, India, 4USLavasa Medical
& Research Center, India, 5Bengaluru Allergy Centre, India.
RATIONALE: Legumes are implicated in IgE mediated food allergy in
different countries. The present study aimed to investigate the effect of
enzymatic hydrolysis on allergenicityof kidney bean, black gramandpeanut.
METHODS: The extracts were subjected to sequential enzymatic treat-
ment by Alcalase and flavorzyme. The allergenicity of hydrolysates were
determined by ELISA and SPT. Allergenic potential of legume proteins
were assessed by ELISA inhibition and basophil histamine release.
RESULTS: Hydrolysis reduced the number of IgE binding fractions of the
three legumes- kidney bean, black gram and peanut. Specific IgE binding
was reduced 62%, 87% and 92% in the hyrolysates of kidney bean, black
gram and peanut, respectively after 8 h of hydrolysis (p<0.01). Alcalase
treatment alone increased the IC50 value of kidney bean, black gram and
peanut to 4, 8.5 and 123 folds respectively. Sequential hydrolysis using
both alcalase and flavourzyme increased the IC50 of the kidney bean, black
gram and peanut to 25, 368 and 393 folds respectively further as compared
to single enzyme treatment. Basophil histamine release showed reduced
allergenicity of the hydrolyzed proteins. Significant reduction in the
biopotency of hydrolyzed protein of kidney bean, black gram and peanut
was observed in SPT where 6/13 kidney bean sensitive individuals, 8/13
black gram sensitive individuals and 9/15 peanut sensitive individuals were
found SPT negative to their respective hydrolysates.
CONCLUSIONS: Sequential hydrolysis by alcalase and flavourzyme
reduced the allergenicity of legume proteins. The method has potential to
reduce allergenicity of food extracts.
68 Comparative Analysis of IgE Binding Proteins in GM and Non-GM Rice Varieties Using Atopic Patients Sera
Chandni Mathur1, P. C Kathuria2, Shakuntala Lavasa3, Pushpa Dahiya4,
Anand Singh, PhD5; 1Institute of Genomics & Integrative Biology, Delhi,
India, 2National Allergy Centre, India, 3USLavasa Medical & Research
Center, India, 4M.D University, India, 5Institute of Genomics and Integra-
tive Biology, Delhi, India.
RATIONALE: Food allergy is recognized as IgE-mediated immune
response. Safety issues regarding foods derived from genetically modified
(GM) plants are central to their acceptance into the food supply and its
commercial production.
METHODS: Three GM rice varieties containing cry protein Cry 1ab, Cry
1ac and Cry 1c along with their native variety were selected for the present
investigation. All the seed powders were processed for protein extraction
and estimation. Extracts were subjected to SDS-PAGE for protein profil-
ing. Simulated Gastric Fluid Digestion (SGF) was also done for all seed
powders. IgE binding protein bands in rice were observed through
Immunoblot with sera of allergic patients. Bioinformatic analysis of above
transgenic proteins across databases was also performed.
RESULTS: Protein content in non-GM rice was 0.9 mg/ml while in GM
rice varieties it varied from 0.95 – 1.2 mg/ml. Proteins fractionated in 11
-14 bands in all varieties within a range of 9-81 kD.Native rice andGM rice
with Cry 1Ab subjected to SGF showed a very faint appearance of smear
around 9kD. Rice variety with Cry 1Ab had faint proteins around 27 and 47
kD. IgE binding protein bands observed in rice varieties were aggregating
around 37-47 kD. Comparative analysis across different databases yielded
that cry proteins used in GM Rice showed sequence identity of < 27 % .
CONCLUSIONS: It is suspected that no major detectable difference was
observed in GM and non-GM rice varieties.
69 Analysis of the Association Between Food Allergy andSensitization to Ltp and Profilin
Felicia Berroa1, M Jose Goikoetxea, PhD, MD2, Marta M.
Ferrer, MD, PhD, FAAAAI2, Paula Cabrera-Freitag, MD, PhD3, Gracia
Javaloyes, MD, PhD1, Maria L Sanz, MD, PhD1, Gabriel Gastaminza,
MD, PhD1; 1Department of Allergy, Clinica Universidad de Navarra,
Spain, 2Department of Allergy, Clinica Universidad de Navarra, Pam-
plona, Spain, 3Clinica Universidad de Navarra, Pamplona, Spain.
RATIONALE: LTP and profilin panallergens are associated with food
allergy (FA). The aim of our study was to evaluate the association of
suffering oral allergy syndrome (OAS) and systemic symptoms (SS)
according to sensitization to LTP or profilin.
METHODS: Retrospective analysis of 238 patients in whom skin prick
test (SPT) to palm profilin and peach LTP(ALK-Abell�o, Madrid, Spain)
was performed. Specific IgE rPhl p 12 and rPru p 3 were analyzed by
ImmunoCAP (Phadia, Sweden) from serum obtained on the day of
consultation. The association to present OAS or SS after ingestion of
plant-food was studied by Chi-square and Odds Ratio(OR) calculation.
RESULTS: Out of 238 patients, 72 (30%) had FA: 36 presented OAS, 20
patients SS and 16 OAS and SS. Eighty-three patients had positve SPT to
profilin and/or LTP (42 profilin, 29 LTP and 12 both panallergens).Skin
sensitization to LTP was associated with: OAS (OR: 3.279 [1.595-6.741];
p50.001) and SS (OR: 5.664[2.598-12.349]; p<0.001). Neither LTP-SPTnor rPru p 3 were associated with OAS without SS. A positive result to
rPrup3 by ImmunoCAP was associated with OAS, only in those patients
suffering also from SS. ImmunoCAP (OR: 2,568 [1,254-5,258]; p50.008)
but not SPT (OR: 2.206[1.121-4.342]; p50.020) to profilin were associ-
ated with OAS without SS.
CONCLUSIONS: Having positive SPTand/or ImmunoCAP is associated
with having systemic symptoms with or without OAS. When OAS is
present without systemic symptoms is associated only with ImmunoCAP
to rPhl p 12.
70 Structure and Function of the Peanut Panallergen Ara h 8Barry K. Hurlburt, PhD1, Lesa Celeste, PhD2, Karolina A.
Majorek3, Jane McBride4, Soheila J. Maleki, PhD1, Wladek
Minor, PhD3, Maksymilian Chruszcz, PhD5; 1USDA-ARS-SRRC, New
Orleans, LA, 2university of south carolina, columbia, SC, 3University of
Virginia, Charlottesville, VA, 4usda-ars-srrc, new orleans, LA, 5University
of South Carolina, Columbia, SC.
RATIONALE: Ara h 8 is hypothesized to be the panallergen responsible
for oral allergy syndrome between birch pollen and peanut. In addition, it
may also be a signaling molecule carrier. We sought to investigate these
notions with structural, and biochemical characterization.
METHODS: Recombinant Ara h 8 was expressed in E. coli and purified.
The structurewas determined using X-ray crystallography. Ligand binding
was tested by displacement of bound fluorescent 8-anilo-1-naphthalenesul-
fonic acid.
RESULTS: Purification by a combination of heat treatment, ammonium
sulfate precipitation and hydrophobic interaction chromatography yielded>95%pureAra h 8.The crystal structurewas solvedbymolecular replacement
and refined at 1.7�A resolution. The overall fold ofAra h 8was very similar to
Bet v 1, as well as Bet v 1 homologs Api g 1 (celery), Gly m 4 (soy), and Pru
av 1 (cherry). Physiologically-relevant ligands have been identified.
CONCLUSIONS: Our results support both the idea that Ara h 8 could be
contributing to oral allergy syndrome between birch pollen and peanut, and
the hypothesis that Bet v 1-like proteins could be transporters of signaling
molecules important for plant development.