Analysis of GST Allergen Cross-Reactivity in a North American Population for Molecular Diagnosis

Abstract

M O N D A Y 603 The Major Allergens of Birch Pollen and Cow Milk, Bet v 1 and Bos d 5, Are Structurally Related to Human Lipocalin 2, Enabling Them to Manipulate T-Helper Cells Depending on Their Load with Siderophore-Bound Iron Erika Jensen-Jarolim, MD, Cristina Gomez-Casado, PhD, Ass.Prof., Luis F. F. Pacios, PhD, Prof., Gerlinde Hofstetter, MSc, BSc, Nadine Mothes-Luksch, MD, Georg A. Roth, MD, Josef Singer, MD, PhD, Araceli Diaz-Perales, PhD, Prof, Franziska Roth-Walter, PhD, Ass.Prof.; Comparative Medicine, Messerli Research Institute of the University of Veterinary Medicine Vienna, Medical University Vienna and University Vienna, Austria, Institute for Pathophysiology and Allergy Research, Center of Pathophysiology, Infectiology and Immunology, Medical University of Vienna, Austria, Vienna, Austria, Biotechnology Department, Center for Plant Biotechnology and Genomics, Madrid, Spain, Biotechnology Department, Center for Plant Biotechnology and Genomics, Technical University of Madrid, Madrid, Spain, Comparative Medicine, Messerli Research Institute of the University of Veterinary Medicine Vienna, Medical University Vienna and University Vienna, Vienna, Austria, AllergyCare, Molecular Diagnostics and Study Center, Vienna, Austria, Department of Anesthesiology, General Intensive Care and Pain Medicine, Medical University of Vienna, Vienna, Austria. RATIONALE: Human lipocalin 2, LCN2, is highly expressed in encounter sites of allergens, as lung and gut. It has immuno-regulatory properties when carrying iron via siderophores (holo) or not (apo). We investigated whether major allergens of birch pollen and milk, Bet v 1 and Bos d 5, might structurally and biologically interfere with LCN2. METHODS: Structural comparison to LCN2 was performed using FATCATflex, CE algorithm and TM-Align methods. Ligand binding was analyzed with AutoDock Vina. Iron-binding was determined by Prussian blue staining. Activated human PBMCs were stimulated for 18h with apoor holo-allergens, or controls. Subsequently, cells and supernatants were analysed by flow cytometry and for their cytokine-content. RESULTS: Both tested allergens shared great structural homology to LCN2. Thus, besides Bos d 5, we could also classify Bet v 1 as a lipocalinlike protein. Both allergens were capable of binding iron via siderophores. When incubated with PBMCs, only the apo-forms of the allergens, but not the holo-forms, were able to promote CD4+ cells and the secretion of IL13. CONCLUSIONS: We conclude that Bet v 1 and Bos d 5 not only structurally mimic human LCN2, but also functionally by their ability to bind iron via siderophores. The apo-forms promote Th2 cells, whereas the holo-forms appear to be immunosuppressive. These results provide for the first time a functional understanding on the principle of allergenicity of major allergens from entirely independent sources, like birch and milk. Supported by Austrian Science Fund FWF F4606-B19,W1205-B09, BES2010-03462 (FPI-Programm, Spanish Government (MICINN/MINECO).