Current Technologies for Complex Glycoproteomics and Their Applications to Biology/Disease-Driven Glycoproteomics

Abstract

Glycoproteomics is an important recent advance in the field of glycoscience. In glycomics, glycan structures are comprehensively analyzed after glycans are released from glycoproteins. However, a major limitation of glycomics is the lack of insight into glycoprotein functions. The Biology/Disease-driven Human Proteome Project has a particular focus on biological and medical applications. Glycoproteomics technologies aimed at obtaining a comprehensive understanding of intact glycoproteins, i.e., the kind of glycan structures that are attached to particular amino acids and proteins, have been developed. This Review focuses on the recent progress of the technologies and their applications. First, the methods for large-scale identification of both N- and O-glycosylated proteins are summarized. Next, the progress of analytical methods for intact glycopeptides is outlined. MS/MS-based methods were developed for improving the sensitivity and speed of the mass spectrometer, in parallel with the software for complex spectrum assignment. In addition, a unique approach to identify intact glycopeptides using MS1-based accurate masses is introduced. Finally, as an advance of glycomics, two approaches to provide the spatial distribution of glycans in cells are described, i.e., MS imaging and lectin microarray. These methods allow rapid glycomic profiling of different types of biological samples and thus facilitate glycoproteomics.

Characterization of intact glycopeptides reveals the impact of culture media on site‐specific glycosylation of EPO‐Fc fusion protein generated by CHO‐GS cells

“…Intact glycopeptide profiling characterizing the site‐specific glycans has emerged as an alternative and complementary glycan analysis tool in recent years. Its application includes biomarker discovery, disease diagnosis and biopharmaceutical product quality control (Bora de Oliveira, Spencer, Barton, & Agarwal, ; Kolarich, Jensen, Altmann, & Packer, ; Moh, Lin, Thaysen‐Andersen, & Packer, ; Narimatsu et al, ). Here we adopted this glycoproteomic method for evaluating the effects of different media compositions on the glycan profile of a recombinant protein, EPO‐Fc, secreted from CHO cells.…”

Toward Completion of the Human Proteome Parts List: Progress Uncovering Proteins That Are Missing or Have Unknown Function and Developing Analytical Methods

“…The Japanese colleagues have been pioneers in glycoproteomics and leaders of the HPP efforts in this domain. Narimatsu et al 34 extensively review current technologies, including improved sensitivity and speed of mass spectrometry, new software for complex spectrum assignment, a unique approach to identify intact glycopeptides using MS1-based accurate masses, and spatial distribution within cells using MS imaging and lectin microarray. There are many applications to biology and disease.…”

Discovery of Pancreatic Ductal Adenocarcinoma-Related Aberrant Glycosylations: A Multilateral Approach of Lectin Microarray-Based Tissue Glycomic Profiling With Public Transcriptomic Datasets

“…Accordingly, glycoproteins with disease-relevant glycosylation changes are fascinating targets for the development of novel biomarkers and therapeutic agents (4,5). To efficiently accelerate this development, a reliable strategy is necessary for the analysis of glycosylation alterations occurring on a specific glycoprotein during disease progression (6,7).…”

“…Lectin microarray was developed as a simple and reliable method for glycan analysis (8,9). This technology allows researchers to obtain global glycomic profiles of N-and O-glycoproteins in biological samples, and these profiles are presented as the signal patterns of interactions between multiple lectins and carbohydrates (6,10,11). Taking advantage of the high sensitivity, this technology is fit to meet the aforementioned demands, and thus has been applied to spatial analysis of tissue sections to compare the glycomic profiles of different regions such as tumor and non-tumor regions (12)(13)(14).…”