Synthetic biology E. coli strain development for expression of functional human GPCRs with roles in human disease

Project Summary Abstract G protein coupled receptorsGPCRsmediate most of our bodyandapos s physiological responses to hormonesneurotransmitters and environmental stimulantsThey are trans membrane proteins that are expressed in every type of cell in the body where their function is to detect molecules outside the cell and transmit signals across the membrane to activate cellular responsesThese proteins are involved in many diseases and present a wide range of opportunities as therapeutic targets in areas including diabetescancercentral nervous system disordersinflammationand painGPCRs have been and will remain among the most important targets of drug discovery in the pharmaceutical industryIn order to have sufficient and functional GPCR proteins for drug screeningan efficient heterologous protein expression system is requiredThe current heterologous expression systems using insect cells or mammalian cells typically result in low yields and require lengthy molecular biology effortsEcoli is a very attractive protein expression system which offers flexibility in genetic manipulationrapid growthease of scaling up and very low costHowevercurrent Ecoli strains are limited in their ability to produce fully functional GPCR proteinsIn this projectPrimordial Genetics aims to develop a novel Ecoli strain for producing functional GPCRs by a patented synthetic biology technology called Function GeneratorThis technology has previously been applied to Ecoli to generate functionality that cannot be created by classical genetic approachesWe propose to screen our combinatorial expression libraries to discover all the genes involved in improved expression of GPCRsThe expression of six well known GPCRs will be validated by biochemical assays and function assaysAt the end of this Phase I projectwe will have demonstrated that for each target GPCR we have at least one FG gene that enables high level expression of the soluble GPCROur Phase II project will comprise further strain development by stacking the trait genes to have a robust Ecoli strain that is universally useful for GPCR productionThe Ecoli production host can be licensed to pharmaceutical companies and can be used to supply the market with GPCRs for functional screeningThis endeavor not only bears great business prospects but also provides the scientific community with valuable Randamp D resources Project Narrative The principal aim of this project is to apply a novel synthetic biology approach to develop protein expression strains of the laboratory bacterium Escherichia coli that are suitable for producing functional G protein coupled receptorGPCRproteinsGPCRs are diverse membrane proteins that are ubiquitous in the human bodyThey are involved in many physiological processes as well as many diseases and have been the most important drug targets in modern medicineEfficient production of milligram quantities of functional GPCR is currently very difficultAlthough Ecoli is widely used for protein productionexpressing functional GPCRs in Ecoli is still challengingThe novel Ecoli expression host to be developed in this project will significantly accelerate the production of GPCRs and drug development work with this important class of human proteinsThis project will not only have a broad impact in drug discovery across many diseases but also will provide the scientific community a valuable resource for structural biology research