PMID- 34671372
OWN - NLM
STAT- PubMed-not-MEDLINE
LR  - 20231107
IS  - 1664-462X (Print)
IS  - 1664-462X (Electronic)
IS  - 1664-462X (Linking)
VI  - 12
DP  - 2021
TI  - The Major Peanut Allergen Ara h 2 Produced in Nicotiana benthamiana Contains 
      Hydroxyprolines and Is a Viable Alternative to the E. Coli Product in Allergy 
      Diagnosis.
PG  - 723363
LID - 10.3389/fpls.2021.723363 [doi]
LID - 723363
AB  - Peanut allergy is a potentially life-threatening disease that is mediated by 
      allergen-specific immunoglobulin E (IgE) antibodies. The major peanut allergen 
      Ara h 2, a 2S albumin seed storage protein, is one of the most dangerous and 
      potent plant allergens. Ara h 2 is posttranslationally modified to harbor four 
      disulfide bridges and three hydroxyprolines. These hydroxyproline residues are 
      required for optimal IgE-binding to the DPYSP(OH)S motifs representing an 
      immunodominant IgE epitope. So far, recombinant Ara h 2 has been produced in 
      Escherichia coli, Lactococcus lactis, Trichoplusia ni insect cell, and 
      Chlamydomonas reinhardtii chloroplast expression systems, which were all 
      incapable of proline hydroxylation. However, molecular diagnosis of peanut 
      allergy is performed using either natural or E. coli-produced major peanut 
      allergens. As IgE from the majority of patients is directed to Ara h 2, it is of 
      great importance that the recombinant Ara h 2 harbors all of its eukaryotic 
      posttranslational modifications. We produced hydroxyproline-containing and 
      correctly folded Ara h 2 in the endoplasmic reticulum of leaf cells of Nicotiana 
      benthamiana plants, using the plant virus-based magnICON((R)) transient expression 
      system with a yield of 200 mg/kg fresh biomass. To compare prokaryotic with 
      eukaryotic expression methods, Ara h 2 was expressed in E. coli together with the 
      disulfide-bond isomerase DsbC and thus harbored disulfide bridges but no 
      hydroxyprolines. The recombinant allergens from N. benthamiana and E. coli were 
      characterized and compared to the natural Ara h 2 isolated from roasted peanuts. 
      Natural Ara h 2 outperformed both recombinant proteins in IgE-binding and 
      activation of basophils via IgE cross-linking, the latter indicating the potency 
      of the allergen. Interestingly, significantly more efficient IgE cross-linking by 
      the N. benthamiana-produced allergen was observed in comparison to the one 
      induced by the E. coli product. Ara h 2 from N. benthamiana plants displayed a 
      higher similarity to the natural allergen in terms of basophil activation due to 
      the presence of hydroxyproline residues, supporting so far published data on 
      their contribution to the immunodominant IgE epitope. Our study advocates the use 
      of N. benthamiana plants instead of prokaryotic expression hosts for the 
      production of the major peanut allergen Ara h 2.
CI  - Copyright (c) 2021 Uzulmez, Kalic, Mayr, Lengger, Tscheppe, Radauer, Hafner, Hemmer 
      and Breiteneder.
FAU - Uzulmez, Oyku
AU  - Uzulmez O
AD  - Institute of Pathophysiology and Allergy Research, Medical University of Vienna, 
      Vienna, Austria.
FAU - Kalic, Tanja
AU  - Kalic T
AD  - Institute of Pathophysiology and Allergy Research, Medical University of Vienna, 
      Vienna, Austria.
AD  - Department of Dermatology, University Hospital St. Polten, Karl Landsteiner 
      University of Health Sciences, St. Polten, Austria.
FAU - Mayr, Vanessa
AU  - Mayr V
AD  - Institute of Pathophysiology and Allergy Research, Medical University of Vienna, 
      Vienna, Austria.
FAU - Lengger, Nina
AU  - Lengger N
AD  - Institute of Pathophysiology and Allergy Research, Medical University of Vienna, 
      Vienna, Austria.
FAU - Tscheppe, Angelika
AU  - Tscheppe A
AD  - Institute of Pathophysiology and Allergy Research, Medical University of Vienna, 
      Vienna, Austria.
FAU - Radauer, Christian
AU  - Radauer C
AD  - Institute of Pathophysiology and Allergy Research, Medical University of Vienna, 
      Vienna, Austria.
FAU - Hafner, Christine
AU  - Hafner C
AD  - Department of Dermatology, University Hospital St. Polten, Karl Landsteiner 
      University of Health Sciences, St. Polten, Austria.
AD  - Karl Landsteiner Institute for Dermatological Research, St. Polten, Austria.
FAU - Hemmer, Wolfgang
AU  - Hemmer W
AD  - FAZ - Floridsdorf Allergy Center, Vienna, Austria.
FAU - Breiteneder, Heimo
AU  - Breiteneder H
AD  - Institute of Pathophysiology and Allergy Research, Medical University of Vienna, 
      Vienna, Austria.
LA  - eng
GR  - P 30936/FWF_/Austrian Science Fund FWF/Austria
PT  - Journal Article
DEP - 20211004
PL  - Switzerland
TA  - Front Plant Sci
JT  - Frontiers in plant science
JID - 101568200
PMC - PMC8522509
OTO - NOTNLM
OT  - Ara h 2
OT  - Nicotiana benthamiana
OT  - PTM
OT  - hydroxyproline
OT  - peanut allergy
OT  - transient expression
COIS- The authors declare that the research was conducted in the absence of any 
      commercial or financial relationships that could be construed as a potential 
      conflict of interest.
EDAT- 2021/10/22 06:00
MHDA- 2021/10/22 06:01
PMCR- 2021/01/01
CRDT- 2021/10/21 06:08
PHST- 2021/06/10 00:00 [received]
PHST- 2021/08/17 00:00 [accepted]
PHST- 2021/10/21 06:08 [entrez]
PHST- 2021/10/22 06:00 [pubmed]
PHST- 2021/10/22 06:01 [medline]
PHST- 2021/01/01 00:00 [pmc-release]
AID - 10.3389/fpls.2021.723363 [doi]
PST - epublish
SO  - Front Plant Sci. 2021 Oct 4;12:723363. doi: 10.3389/fpls.2021.723363. eCollection 
      2021.