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Team:Brown/Project
From 2009.igem.org
A Synthetic Approach to Treating Allergic Rhinitis: Engineering Staphyloccocus Epidermidis to Secrete High-Affinity Histamine Binding Protein in Response to Elevated Levels of Histamine during an Allergic Attack
Project Abstract
Brown University’s 2009 iGEM Team presents an exciting new approach to treating nasal allergies through Allergene: a synthetically engineered, self-regulating drug factory in the nose. This revolutionary new product provides a much-needed alternative to current antihistamines by directly sequestering the histamine released in an allergic response. In order to do this, Allergene makes use of the unique histamine binding protein rEV131, native to the tick Rhipicephalus appendiculatus. By taking advantage of rEV131's high binding affinity for histamine, Allergene effectively eliminates both the symptoms and side effects associated with allergies and their treatments.
Rather than presenting a system that passively sequesters histamine; however, Allergene goes one step further to providing patients with much-needed allergy relief. Activation of the product only occurs in the explicit event of an actual allergic response. This ingenious mechanism is made possible through the novel engineering of a histamine receptor in prokaryotes, a grand undertaking that had never before been accomplished. By re-designing pre-existing prokaryotic chemoreceptors to bind histamine rather than their wild-type ligands, Allergene acquired its most impressive feature yet: the "Histamine Sensor". Site-directed mutagenesis on the ligand binding pockets of two particular prokaryotic chemoreceptors, Ribose Binding Protein and Tar (normally responsive to Aspartate)was performed to create this novel sensor. Through histamine detection, Allergene’s unique histamine sequestering system is engineered to function only when histamine levels are markedly high. This efficiently timed system is therefore completely self-regulating.
Allergene is introduced to its host patients by capitalizing on the endogenous existence of bacterium Staphylococcus epidermidis in human nasal flora. This naturally present organism is the perfect vehicle for delivery; its rapid production and secretion of Allergene’s genetic constructs effectively implement the system’s histamine sensing and sequestering capabilities in human hosts. Secretion is accomplished through the attachment of a signal peptide sequence specific to S. epidermidis, a clear and concise method by which to deliver the ready-to-use drug. In order to eliminate any safety concerns associated with the utilization of S. epidermidis, special care has additionally been taken to engineer the product’s accessory kill switch mechanism. Capitalizing on the cells' abilities to sense the growth of their own populations, a DNA gyrase poison responsible for triggering cell death (CCDB)is neatly placed under the quorum or population sensing promoter Agr, which normally functions in hazardous biofilm formation. Named the “Quorum Sensor”, this thoughtful, additional feature prompts Allergene’s swift response in the unfavorable event of over-cell-proliferation.
