October 5, 2023 11:00 am
October 5, 2023 12:00 pm
Chemical Tools to Study Biological Systems
Monika Raj
Associate Professor of Chemistry
Emory University
The human proteome is extremely complex, comprising > 10,000 proteins and 100 times proteoforms for each gene product. In cancer and other diseases, several new protein variants may result from mutations, fusions and posttranslational modifications (PTMs) that further influence the functions and structure of proteins. This necessitates the identification of proteins and PTMs at a single molecule level in a cell or an organism to understand biological processes, disease analysis and biomarker discovery. Despite the power of protein sequencing in revolutionizing precision medicine diagnostics, there are no efficient single molecule protein sequencing (SMPS) methods to identify several nonreactive amino acids and PTMs at the proteome-wide level. We have developed multiple bioconjugation approaches for the selective labeling of amino acids of poor reactivity1 and mono-methyl lysine,2-3 N-methyl terminus,4 di-methyl lysine and monomethylhistidine posttranslational modifications (PTMs) to fill the present gap in the range of available techniques to sequence and identify proteins and PTMs at the single molecule and single cell level with high sensitivity and high accuracy. The broad utility of these bioconjugation reactions is demonstrated by the conjugation of various affinity probes and fluorophores on amino acids on proteins and PTMs. We showed the utility of our chemical methods in identifying PTMs at the single molecule level by using fluorosequencing SMPS technology. These chemical approaches are broadly applicable to other SMPS technologies and thus have the potential to further our understanding of the role of methylated lysine- and histidine-containing PTMs in regulating various cellular signaling processes and aid in biomarker discovery.complex has demonstrated an unexpected photophysical dependence on the ligand protonation state that we rationalize using density functional theory calculations.