Date of Graduation

5-2025

Document Type

Thesis

Degree Name

Bachelor of Science in Chemistry

Degree Level

Undergraduate

Department

Chemistry & Biochemistry

Advisor/Mentor

Adams, Paul

Committee Member

Hershberger, Margaret

Second Committee Member

Alrubaye, Adnan

Third Committee Member

Lessner, Faith

Abstract

The complexity of the proteome requires strategies that efficiently characterize protein functions. Activity-Based Protein Profiling (ABPP) is a method that identifies enzyme activity by targeting key catalytic residues. This study focused on histidine, a residue involved in enzymatic catalysis, metal coordination, and protein-protein interactions. Despite its importance, histidine-specific chemical probes are limited due to its variable reactivity. This work developed and characterized a histidine-specific probe using a pyrocarbonate-based mechanism. Singleresidue assays confirmed selective reactivity of the probe with histidine over other nucleophilic amino acids. Further testing on enzymatic proteins and lysed cell proteomes showed that the probe modified histidine residues with minimal off-target effects. Liquid chromatography confirmed selective labeling of histidine on the small-molecule scale. These results demonstrate the utility of pyrocarbonate probes for characterization by ABPP. These probes provide a method for studying enzyme activity and protein function, with applications in enzymology, drug discovery, and biomarker identification.

Keywords

proteomics; histidine; pyrocarbonate

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