Mass spectrometry is an extraordinarily powerful bio-analytical technique that has had a profound impact on our molecular understanding of human health and disease. Major advances in mass analyzer technology, gas-phase separations, dissociation techniques, and ionization methods are largely attributed to the central role that mass spectrometry plays in the field of systems biology. While mass spectrometry has evolved over the last century into a highly effective analytical tool, there are still opportunities for new advances, allowing an even more diverse array of biological questions to be addressed. This proposal is centered on the development and characterization of novel, multi-functional, glycan- specific chemical tags for increased detection by spectroscopic (i.e., absolute quantification) and biological mass spectrometry (i.e., relative quantification) methods. Moreover, the establishment of gas- phase separations of glycans to overcome the isomer barrier challenge is essential to fully understanding the underlying biology. The short-term objective of this proposal is to develop and validate a library of novel multi-functional tags and ion mobility (IM) measurements to resolve glycan isomers, with the long-term goal being their application in an Alzheimer?s disease human tissue repository. This project will also develop open-source software for automated glycan analysis to facilitate accurate and high-throughput analysis, including the support of IM data. These N-glycan results will be combined with data on the glycoproteome and proteome, contributing to the ultimate goal elucidating glycans and glycoproteins for a deeper understanding of the role a key protein modification has in the development of AD. Public Description of Proposed Research Mass spectrometry (MS), the science related to the ?weighing of molecules?, has had a profound impact on the study of human health and disease including cancer, heart disease, neural development, and auto-immune diseases. However, this research will leverage front-end chemistries and gas-phase separations for MS-based glycomics/glycoproteomics to dramatically improve the ability of MS to detect, quantify, and resolve glycan isomers extracted from brain tissue proteins. This will address a more diverse array of contemporary biomedical questions including the quantification of diagnostic and prognostic biomarkers. This proposal is centered on the elucidation of a glycan-specific biomarkers and which proteins are modified for understanding the diagnosis and progression of AD. Another major benefit of this research will be to provide new drug targets (e.g., glycoproteins).
This proposal seeks support to develop and apply novel multi-functional tags that will facilitate both global and targeted quantitative mass spectrometric analysis of N-linked glycans with significantly improved limits-of-detection with the ability to resolve isomers using gas-phase separations. This application of technology will help us to understand the role of glycosylation in the development of Alzheimer's disease (AD); glycosylation is a key protein co-translational modification whose functions include protein folding, cell signaling, enzymatic activity, and adhesion. Defining the role of glycosylation in AD patients relative to controls will provide a new level of understanding of how this important modification is involved in the development of AD.
