The influence of recombinant production on the immunologic behavior of birch pollen isoallergens.
BACKGROUND
Allergic reactions towards the birch major pollen allergen Bet v 1 are among the most common causes of spring pollinosis in the temperate climate zone of the Northern hemisphere. Natural Bet v 1 is composed of a complex mixture of different isoforms. Detailed analysis of recombinant Bet v 1 isoforms revealed striking differences in immunologic as well as allergenic properties of the molecules, leading to a classification of Bet v 1 isoforms into high, medium, and low IgE binding proteins. Especially low IgE binding Bet v 1 isoforms have been described as ideal candidates for desensitizing allergic patients with allergen specific immunotherapy (SIT). Since diagnosis and therapy of allergic diseases are highly dependent on recombinant proteins, continuous improvement of protein production is an absolute necessity.


METHODS
Therefore, two different methods for recombinant production of a low IgE binding Bet v 1 isoform were applied; one based on published protocols, the other by implementing latest innovations in protein production. Both batches of Bet v 1.0401 were extensively characterized by an array of physicochemical as well as immunological methods to compare protein primary structure, purity, quantity, folding, aggregation state, thermal stability, and antibody binding capacity.


CONCLUSIONS
The experiments demonstrated that IgE antibody binding properties of recombinant isoallergens can be significantly influenced by the production method directly affecting possible clinical applications of the molecules.
Wallner
Michael
M
Christian Doppler Laboratory for Allergy Diagnosis and Therapy, University of Salzburg, Salzburg, Austria. michael.wallner@sbg.ac.at
Himly
Martin
M
Neubauer
Angela
A
Erler
Anja
A
Hauser
Michael
M
Asam
Claudia
C
Mutschlechner
Sonja
S
Ebner
Christof
C
Briza
Peter
P
Ferreira
Fatima
F
D000485:Allergens / Q000737:chemistry / Q000276:immunology* / Q000302:isolation & purification; D000818:Animals; D029662:Betula / Q000276:immunology*; D002942:Circular Dichroism; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007073:Immunoglobulin E / Q000276:immunology; D008024:Ligands; D008970:Molecular Weight; D010940:Plant Proteins / Q000737:chemistry / Q000302:isolation & purification; D011058:Pollen / Q000276:immunology*; D011489:Protein Denaturation; D017510:Protein Folding; D020673:Protein Renaturation; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D051381:Rats; D011994:Recombinant Proteins / Q000096:biosynthesis*; D012995:Solubility
D000485:Allergens; D008024:Ligands; D010940:Plant Proteins; D011994:Recombinant Proteins; D007073:Immunoglobulin E