PMID- 15593144
OWN - NLM
STAT- MEDLINE
DCOM- 20050808
LR  - 20061115
IS  - 1615-9853 (Print)
IS  - 1615-9853 (Linking)
VI  - 5
IP  - 1
DP  - 2005 Jan
TI  - Novel isoforms of Pru av 1 with diverging immunoglobulin E binding properties 
      identified by a synergistic combination of molecular biology and proteomics.
PG  - 282-9
AB  - Birch pollen-related food allergies are mainly associated to Bet v 1. Little is 
      known about isoforms of Bet v 1 homologous in fruit of the Rosaceae family. We 
      attempted to identify novel isoforms of Pru av 1, the major cherry allergen, at 
      the cDNA and the protein level by a combination of molecular biology and 
      proteomic tools. A cDNA library was screened with patients immunoglobulin E (IgE) 
      and a specific hybridization probe. Edman sequencing, mass spectrometry (MS), and 
      MS/MS were performed after detecting Pru av 1 on 2-D maps by immunoblotting using 
      patients IgE and a monoclonal antibody. Partial amino acid sequences were 
      completed with a polymerase chain reaction (PCR) strategy. The IgE-binding 
      properties of the Pru av 1 spots were analyzed by 2-D blot inhibition. cDNA 
      library analysis revealed a novel Pru av 1 isoform. MS and N-terminal sequencing 
      confirmed the cDNA sequences at the protein level. A series of spots were 
      confirmed as the already known Pru av 1. One spot, exclusively detected with 
      patients sera, was identified as the novel isoform. A partial amino acid sequence 
      detected with MS/MS was completed by PCR-cloning. The 2-D blot inhibition 
      revealed epitope differences between the novel isoform and the previously 
      published Pru av 1. Our data demonstrate that a synergistic combination of 
      molecular biology and proteomics represents a powerful tool for reliable and 
      comprehensive identification of allergen isoforms and variants. The newly 
      identified isoform showed diverging IgE-binding properties and may be relevant 
      for the diagnosis or therapy of cherry allergy.
FAU - Reuter, Andreas
AU  - Reuter A
AD  - Paul-Ehrlich-Institut, Division of Allergology, D-63225 Langen, Germany.
FAU - Fortunato, Donatella
AU  - Fortunato D
FAU - Garoffo, Lorenza Perono
AU  - Garoffo LP
FAU - Napolitano, Lorenzo
AU  - Napolitano L
FAU - Scheurer, Stephan
AU  - Scheurer S
FAU - Giuffrida, Maria G
AU  - Giuffrida MG
FAU - Vieths, Stefan
AU  - Vieths S
FAU - Conti, Amedeo
AU  - Conti A
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PL  - Germany
TA  - Proteomics
JT  - Proteomics
JID - 101092707
RN  - 0 (Allergens)
RN  - 0 (Peptide Library)
RN  - 0 (Plant Proteins)
RN  - 0 (Protein Isoforms)
RN  - 37341-29-0 (Immunoglobulin E)
SB  - IM
MH  - Allergens/genetics/*metabolism
MH  - Amino Acid Sequence
MH  - Electrophoresis, Gel, Two-Dimensional
MH  - Gene Library
MH  - Humans
MH  - Immunoglobulin E/genetics/*metabolism
MH  - Molecular Sequence Data
MH  - Peptide Library
MH  - Plant Proteins/genetics/*metabolism
MH  - Pollen
MH  - Polymerase Chain Reaction
MH  - Protein Binding
MH  - Protein Isoforms/genetics/metabolism
MH  - Proteomics
MH  - Rosaceae/genetics/*immunology
MH  - Spectrometry, Mass, Electrospray Ionization
MH  - Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
EDAT- 2004/12/14 09:00
MHDA- 2005/08/09 09:00
CRDT- 2004/12/14 09:00
PHST- 2004/12/14 09:00 [pubmed]
PHST- 2005/08/09 09:00 [medline]
PHST- 2004/12/14 09:00 [entrez]
AID - 10.1002/pmic.200400874 [doi]
PST - ppublish
SO  - Proteomics. 2005 Jan;5(1):282-9. doi: 10.1002/pmic.200400874.