Modern biomedical research has been profoundly changed by the availability of complete genome sequences and the development of tools such as deep sequencing and microarrays to leverage these sequences for both research and diagnostic applications. The availability of tools to carry out similarly large scale analyses of the whole proteome would be equally revolutionary. Such a system could be used for a broad range of applications, including mapping enzyme substrate specificity (e.g. of kinases, phosphatases, and proteases), immune-repertoire profiling (e.g. determining which specific food allergens an allergic patient has antibodies to), and protein-protein interaction screening. However, to date, the technical challenges of producing and assaying large numbers (tens of thousands) of proteins or peptides have prevented the development of such tools. Here we propose a new technology platform for the programmable synthesis and assay of proteome-scale peptide arrays. We have previously developed a microfluidic platform to produce spectrally encoded beads, which can be used as a substrate for peptide synthesis. Sorting of these beads, based on their codes, into 20 pools, will allow us to perform stepwise peptide synthesis on the beads, resulting in a collection of beads where each spectrally encoded bead is uniquely associated with a peptide sequence. This one-to-one mapping of peptides to beads allows for code-directed, step-by-step synthesis of peptides on beads and later rapid identification of bead-associated peptides by imaging alone. We expect that this technology will result in a powerful new tool for the study of antibody repertoires, enzyme-substrate interactions, and other protein-protein interactions.

Public Health Relevance

Protein interactions with other proteins are a key component of biological systems, and include medically relevant interactions such as antibodies binding to the antigens they recognize. However, we currently have few good tools with which to study them. We are developing a new system to measure the binding of thousands of protein fragments to the proteins that recognize them. This system will be a useful tool in the study of many different biological and medical problems. We will validate the system by determining the particular peanut allergen fragments recognized by the antibodies of patients with peanut allergies.

Agency
National Institute of Health (NIH)
Type
Research Project (R01)
Project #
1R01GM107132-01A1
Application #
8762296
Study Section
Instrumentation and Systems Development Study Section (ISD)
Program Officer
Edmonds, Charles G
Project Start
Project End
Budget Start
Budget End
Support Year
1
Fiscal Year
2014
Total Cost
Indirect Cost
Name
University of California San Francisco
Department
Biochemistry
Type
Schools of Medicine
DUNS #
City
San Francisco
State
CA
Country
United States
Zip Code
94143