Date of Graduation
Summer 2018
Degree
Master of Science in Chemistry
Department
Chemistry and Biochemistry
Committee Chair
Keiichi Yoshimatsu
Abstract
Autocatalytic intramolecular isopeptide bonds have been found in nature in certain gram-positive bacterial pilus structures. Recently, splitting of these domains that are capable of autocatalytic intramolecular isopeptide bond formation have been applied to create stable, selective, bio-orthogonal Catcher/Tag systems. The CnaB2 domain found in the FbaB pilus structure of Streptococcus pyogenes, has yielded the Catcher/Tag, Protein/Peptide systems termed SpyCatchter and SpyTag. Recent study has focused on tag optimization, stability and bio-orthogonality evaluation, along with applications in bioconjugation. I have recombinantly expressed SpyCatcher and SpyTag-fused proteins in E.coli, and conjugated them to fluorescent probes in order for use in fluorescence polarization/depolarization study. Using this system, I have observed the dependence of SpyTag concentration on the formation of the isopeptide bond. I have also been able to track the formation of this bond in real time.
Keywords
spycatcher, spytag, kinetics, fluorescence polarization, isopeptide bond
Subject Categories
Analytical Chemistry | Biochemistry | Biotechnology
Copyright
© Samuel Patricc Kasson
Recommended Citation
Kasson, Samuel Patricc, "Development of Rapid, Homogeneous Assay for Investigating Isopeptide Bond Formation Using Fluorescence Polarization/Depolarization Measurements" (2018). MSU Graduate Theses/Dissertations. 3292.
https://bearworks.missouristate.edu/theses/3292
Open Access
