Hypoallergenic profilin--a new way to identify allergenic determinants.
Plant profilins are important panallergens, contributing to numerous food and pollen allergies. They fulfil essential functions in all eukaryotic cells, including yeast and mammals, making breeding of profilin-deficient crop plants an impossible task. To obtain hypoallergenic profilin-variants, a novel yeast based screening system to select for fully active but hypoallergenic profilins was developed. Despite similar biological functions, plant profilins share only limited amino acid sequence identity to nonplant profilins. Thus, it should be possible to select for hypoallergenic profilin variants that have kept their biological function. These variants were subsequently tested for IgE-binding and their 3D folding was analyzed. As prerequisites, we developed a conditional profilin-deficient yeast mutant strain and made use of a newly-discovered profilin variant from tomato that enabled pre-selection for efficient mutagenesis. After random mutagenesis and transformation of tomato profilin (Lyc e 1) into the designed yeast strain, we were able to screen for functional tomato profilin variants. Testing these variants for IgE binding revealed that two variants showed a strong reduction in IgE binding of four Lyc e 1 sensitized patients and one birch pollen allergic patient. This result, for the first time, provides strong evidence that selection of hypoallergenic Lyc e 1 variants is possible. Furthermore, this knowledge provides the basis for the molecular breeding of hypoallergenic profilin alleles in tomato.
Department of Biology, Friedrich-Alexander-University Erlangen-Nuremberg, Staudtstrasse 5, Erlangen, Germany.
Kathrin E
KE
Paulus
Benedikt
B
Schmid
Doreen
D
Zajic
Alice
A
Schäfer
Vera
V
Mahler
Uwe
U
Sonnewald
D000485:Allergens / Q000737:chemistry / Q000235:genetics* / Q000276:immunology*; D000595:Amino Acid Sequence; D000818:Animals; D052179:Antigens, Plant; D001483:Base Sequence; D018744:DNA, Plant / Q000235:genetics; D000939:Epitopes / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D005512:Food Hypersensitivity / Q000276:immunology / Q000517:prevention & control; D005816:Genetic Complementation Test; D014644:Genetic Variation; D006801:Humans; D007073:Immunoglobulin E / Q000378:metabolism; D066298:In Vitro Techniques; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016296:Mutagenesis; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D051304:Profilins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D012441:Saccharomyces cerevisiae / Q000235:genetics
D000485:Allergens; D052179:Antigens, Plant; D018744:DNA, Plant; D000939:Epitopes; C488367:Lyc e 1 allergen, Lycopersicon esculentum; D010940:Plant Proteins; D051304:Profilins; D011994:Recombinant Proteins; D007073:Immunoglobulin E