Dissertation Title: Tandem Display for Library-on-Library Protein Interactions Screens 

Abstract: 

Proteomics describes the set of all proteins and their expression levels in their biological context. Underlying the expression of every gene in the genome is a rich network of regulating interactions, each modulating the final expression of the encoded proteins. Gene expression defines the cell state, and its identity. In the disease state, expression levels in the system deviate from healthy levels. With a molecular understanding of disease, it becomes possible to identify drug targets that may drive the system back to a healthy state.

Proteomics defines the cell state, yet it is precisely this molecular state that is so hard to measure. Proteins, unlike nucleic acids, are not trivial to sequence and quantify in high throughput. Proteomics measurements are typically performed on mass spectrometers that can only run a handful of samples at a time.

In this thesis we are motivated to resolve the throughput limitations of traditional proteomics measurement by converting proteomics to a next generation sequencing measurement using DNA barcoded antibodies. Since there are not enough high-quality antibodies for whole proteome coverage, we invented a new method for called Tandem Display, a Library-on-Library approach for antibody selection against the whole human proteome simultaneously. Unlike previous approaches, Tandem Display exhibits constant kinetic scaling despite quadratic growth in combinatorial diversity with increasing library complexity.  In this defense we describe progress toward large complexity interaction screens using a model covalent protein framework called the SpyCatcher-SpyTag system and identify orthogonal orthologs in a 400x400 scale screen. Finally, we describe future work to adapt the technique to non-covalent selection and its potential use to validate affinity and specificity of de novo designed protein binders.

Committee members: 
Edward Boyden
Professor of Brain and Cognitive Sciences
MIT

George Church
Robert Winthrop Professor of Genetics
Harvard Medical School

Vishal Tandon
Group Leader - Biological Microsystems
Charles Stark Draper Laboratory