Date of Award
5-19-2023
Degree Name
Doctor of Philosophy (PhD)
Degree Type
Dissertation
Abstract
As the need for protein higher order structure (HOS) characterization approach grows, mass spectrometry (MS)-based structural proteomics tools can be designed to play a role because they have the capacity to measure protein HOS with relatively high resolution and high efficiency. Among these methods, footprinting usually involves chemical labeling of the protein of interest and sequential bottom-up LC−MS/MS analysis to locate the labeled amino acids. This dissertation is focused on footprinting method developments and their applications to characterize therapeutic and disease-related proteins. It is divided into six chapters, among which four chapters are descriptions of original research. The first chapter serves as a general introduction to protein HOS and MS-based structural proteomics tools, paving the way for the research projects that are discussed in later chapters. In chapter 2, the comparison of sensitivity of FPOP−MS and other biophysical approaches for detecting subtle HOS changes of a monoclonal antibody upon thermal stress is reported (chapter 2.1). The application of FPOP−MS to study antibody HOS in different buffers is also discussed (chapter 2.2). Chapter 3 discusses the characterization of a mouse-derived antibody against Marburg virus nucleoprotein. Chapter 4 highlights the development of a fast photochemical oxidation of protein (FPOP)-MS based platform to study protein thermal denaturation, using the lambda repressor protein as an example. Chapter 5 describes the characterization of a novel glycosidase produced by a gut microbiota. Chapter 6 serves as a conclusion to this dissertation, discussing the characteristics and limitations of footprinting methods, proposed future development, and biochemical findings.
Language
English (en)
Chair and Committee
Michael Gross
Recommended Citation
Lin, Yanchun, "Mass Spectrometry-Based Characterization of Protein Higher Order Structure: Method Developments and Applications" (2023). Arts & Sciences Electronic Theses and Dissertations. 2985.
https://openscholarship.wustl.edu/art_sci_etds/2985