Novel Techniques for Characterization of Biomolecular Behavior Across Multiple Biological Scales
Abstract
The development of the central dogma of molecular biology in the late 1950’s unlocked a now booming field of medical innovation centered about the study of biomolecules. Of these, proteins play perhaps the most active role, acting as molecular devices to carry out dynamic processes of cellular function and life maintenance including energy metabolism, DNA replication, cellular signaling, and many others. Historically, proteins have been studied in small subsets; however, there is a recent paradigm shift toward studying proteins and their interactions within complex, physiologically relevant conditions. This shift is driven by advances in both sequencing and computing technologies that enable novel approaches in integrative biology. Here, I present two such techniques that combine computational and experimental methods to elucidate and quantify protein behavior across multiple biological scales. The first is a tool to measure protein-protein interaction (PPI) kinetics. Fluorescence Rolling Correlation Spectroscopy (FRCS) is a novel algorithm and software that applies principles of diffusometry to enable facile characterization of kinetics across the entire dynamic range of typical PPIs [ka£(1e4, 1e6) M−1 ·s −1 , kd£(1e−4, 1e1) s−1 ]. The second is a kinetic model of non-canonical amino acid proteome labeling. I present a system of ordinary differential equations describing Azidohomoalanine (Aha) distribution and selective labeling of nascent proteins in vivofor either enrichment or imaging. This model demonstrates accurate predictions of labeling across multiple tissues and timescales. Each of these tools is presented with open source software and models to enable future work in the study of protein behavior.
Degree
M.Sc.
Advisors
Kinzer-Ursem, Purdue University.
Subject Area
Molecular biology|Agronomy|Atomic physics|Biology|Genetics|Mathematics|Medicine|Physics
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