The central dogma of biology describes fundamentals of molecular components DNA, RNA and proteins. Translation of heritable information from nucleotide sequences into proteins comprises underlying circuitry of all biological kingdoms. At the molecular level protein expression changes represent the primary basis for phenotypic features and physiological diversity among all forms of life. In addition, perturbations in protein expressions are root causes for many diseases. Recent technologies facilitate genome wide quantitation of DNA or RNA in a day, and have become commonplace throughout biology and medicine. An analogous method for rapid, inexpensive global scale protein quantitation is lagging. This proposal executes proof of principle for seed components of an innovative proteome array technology that will measure protein levels in a rapid, simultaneous, cost-effective, multiplexed and scalable format. The innovative method ultimately allows quantitative proteome-wide protein determinations in a single assay, probing complex cellular mixtures, paralleling current genomics throughput. This technology has impact for proteome scale queries: 1) as a research tool to study molecular function of biological systems and 2) identifying changes in protein expressions during disease. We also envision wider utility including proteome-wide-association population analyses and clinical diagnostic applications. We foresee no limit with respect to target cell type, tissue or species, and high reproducibility, linear range and sensitivity. This technology may also be used to quantitate protein modifications, like phosphorylation, glycosylation, methylation, acetylation or amino acid polymorphism. Methods: This proposal synergizes two established assay principles, facilitating a new technology for quantitative whole proteome assay. We will engineer subsets of protein detection reagents, and test specific identification of affinity paired protein targets within a complex pool. Objectives: 1) Establish proof of principle using a novel barcoding technology for multiplex protein detection. 2) Generating a plurality of assays to demonstrate a path towards scalability for whole proteome coverage. Future goals: a) After proof of concept is executed, decoding sufficient protein detection reagents will generate a new tool to measure all ~20,000 human proteins simultaneously. b) Subsequent assay deployment refining component specificities will be developed to simultaneously quantitate proteomes of other species. Agency relevance: This proposal defines a proof of principle and subsequently proof of scalability, for a novel technology that will facilitate simultaneous whole proteome quantitation. Once in practice, this method will have wide reaching impact for study of protein changes in complex biological systems. Proteome analysis will accelerate molecular discovery underlying pathophysiology.
Relevance to public health: This application executes proof of principle for seed components underlying a novel whole- proteome-quantitation technology. Pending successful outcomes, wide biological impact is envisioned: identifying protein changes to explain biological function or molecular causes for disease.
