Combination of High-Resolution Multistage Ion Mobility and Tandem MS with High Energy of Activation to Resolve the Structure of Complex Chemoenzymatically Synthesized Glycans

Abstract Simple and complex carbohydrates (glycans) have long been known to play major metabolic, structural and physical roles in biological systems. Targeted microbial binding to host glycans has also been studied for decades. But such biological roles can only explain some of the remarkable complexity and organismal diversity of glycans in nature. Reviewing the subject about two decades ago, one could find very few clear-cut instances of glycan-recognition-specific biological roles of glycans that were of intrinsic value to the organism expressing them. In striking contrast there is now a profusion of examples, such that this updated review cannot be comprehensive. Instead, a historical overview is presented, broad principles outlined and a few examples cited, representing diverse types of roles, mediated by various glycan classes, in different evolutionary lineages. What remains unchanged is the fact that while all theories regarding biological roles of glycans are supported by compelling evidence, exceptions to each can be found. In retrospect, this is not surprising. Complex and diverse glycans appear to be ubiquitous to all cells in nature, and essential to all life forms. Thus, >3 billion years of evolution consistently generated organisms that use these molecules for many key biological roles, even while sometimes coopting them for minor functions. In this respect, glycans are no different from other major macromolecular building blocks of life (nucleic acids, proteins and lipids), simply more rapidly evolving and complex. It is time for the diverse functional roles of glycans to be fully incorporated into the mainstream of biological sciences.

The glycome describes the complete repertoire of glycoconjugates composed of carbohydrate chains, or glycans, that are covalently linked to lipid or protein molecules. Glycoconjugates are formed through a process called glycosylation and can differ in their glycan sequences, the connections between them and their length. Glycoconjugate synthesis is a dynamic process that depends on the local milieu of enzymes, sugar precursors and organelle structures as well as the cell types involved and cellular signals. Studies of rare genetic disorders that affect glycosylation first highlighted the biological importance of the glycome, and technological advances have improved our understanding of its heterogeneity and complexity. Researchers can now routinely assess how the secreted and cell-surface glycomes reflect overall cellular status in health and disease. In fact, changes in glycosylation can modulate inflammatory responses, enable viral immune escape, promote cancer cell metastasis or regulate apoptosis; the composition of the glycome also affects kidney function in health and disease. New insights into the structure and function of the glycome can now be applied to therapy development and could improve our ability to fine-tune immunological responses and inflammation, optimize the performance of therapeutic antibodies and boost immune responses to cancer. These examples illustrate the potential of the emerging field of 'glycomedicine'.

Summary: The ProteoWizard software project provides a modular and extensible set of open-source, cross-platform tools and libraries. The tools perform proteomics data analyses; the libraries enable rapid tool creation by providing a robust, pluggable development framework that simplifies and unifies data file access, and performs standard proteomics and LCMS dataset computations. The library contains readers and writers of the mzML data format, which has been written using modern C++ techniques and design principles and supports a variety of platforms with native compilers. The software has been specifically released under the Apache v2 license to ensure it can be used in both academic and commercial projects. In addition to the library, we also introduce a rapidly growing set of companion tools whose implementation helps to illustrate the simplicity of developing applications on top of the ProteoWizard library. Availability: Cross-platform software that compiles using native compilers (i.e. GCC on Linux, MSVC on Windows and XCode on OSX) is available for download free of charge, at http://proteowizard.sourceforge.net. This website also provides code examples, and documentation. It is our hope the ProteoWizard project will become a standard platform for proteomics development; consequently, code use, contribution and further development are strongly encouraged. Contact: darren@proteowizard.org; parag@ucla.edu Supplementary information: Supplementary data are available at Bioinformatics online.