Congruent Strategies for Carbohydrate Sequencing. 3. OSCAR:  An Algorithm for Assigning Oligosaccharide Topology from MSⁿ Data

Abstract

This is the third in a sequence of reports devoted to the development of congruent strategies for carbohydrate sequencing. Two previous reports outlined the strategies for observing structural detail from MS n data and introduced tools that compile, search, and compare fragment spectra in a bottomup approach to oligosaccharide sequencing. In this third report, we introduce the operational details of an algorithm that we define as the Oligosaccharide Subtree Constraint Algorithm (OSCAR). This algorithm assimilates analyst-selected MS n ion fragmentation pathways into oligosaccharide topology (branching and linkage) using what may be considered a top-down sequencing strategy. Guided by a series of logical constraints, this de novo algorithm provides molecular topology without presumed biosynthetic constraints or external comparisons. In this introductory study, OSCAR is applied to a series of permethylated oligomers and isomeric glycans, and topologies are assigned in a few hundredths of a second.The two preceding reports in this series introduced methodologies for determining the structural details of carbohydrates from MS n data 1 and described tools that extract further details through spectral comparisons against known oligosaccharide fragments. 2 Glycobiology in general has been a fertile area for the development of bioinformatics tools, 3 and a variety of computer programs are now available to support carbohydrate analysis using mass spectrometry data. The web-based tool GlycoMod 4 accepts a glycan MS mass and returns a list of possible compositions, using literature-derived constraints to limit its output. In contrast, StrOligo 5 , 6 examines oligosaccharide MS 2 spectra to propose a set of candidate structures restricted by biosynthetic constraints.The candidates are fragmented in silico, and the resulting simulated spectra are ranked against the experimental MS 2 spectrum. Similarly, GlycosidIQ 7 compares an experimental MS 2 spectrum against simulated spectra generated from the contents of GlycoSuiteDB, 8 a curated database of known structures, to produce a ranked list of candidate glycans. In the catalog library method, 9 , 10 a catalog contains the characteristic fragmentation patterns of substructures isolated from a library of known oligosaccharides. Total structure assignment is accomplished by matching observed fragmentation patterns with the catalog motif entries. NIH Public AccessThe web-based Saccharide Topology Analysis Tool (STAT) accepts a native glycan mass as input, infers the glycan's possible compositions over a wide range of monosaccharide residues, and generates a candidate set of all possible branching topologies. 11 , 12 STAT then accepts a list of ion masses extracted from the MS n data tree, where each ion must form a connected substructure within the candidate topologies. Finally, a ranked list of branching topologies is generated. STAT supports native glycans, requires manual intervention to resolve ambiguous ion compositions, and restricts the branch points for N-linked ...

Carbohydrate Structural Isomers Analyzed by Sequential Mass Spectrometry

“…In these examples, we will use many of the concepts and tools previously described. 7,8,15,[26][27][28] All structures were derived de novo with input data from a single instrument.…”

“…Structures, whether new or previously described, and their pathways were entered into GlySpy, a set of tools for data handling, complete with the glycan topology assignment algorithm, OSCAR. 28 Ion m/z fragmentation pathways used for OSCAR were selected manually by the analyst. OSCAR was then used to determine which of the isomeric structures were consistent with the fragmentation pathways.…”

“…25 In our experience, spectral reproducibility has been excellent, 26 and this very feature has provided an opportunity to build a searchable fragment library 27 and derive glycan structure with supporting tools. 28 It has been effectively argued that the transfer of energy by gas-phase collisions maybe too broad to define the subtle isomeric features within carbohydrate structures. For this reason, an alternative excitation strategy, infrared multiple-photon dissociation (IR-MPD), 29 was evaluated with a series of glucose disaccharide isomers.…”

A glycomics platform for the analysis of permethylated oligosaccharide alditols

“…Online derivatization coupled with LC/MS of permethylated oligosaccharide alditols has obvious advantages, and we are in the process of adapting online derivatization to the LC/MS platform. Reinhold et al have recently reported the complete assignment of oligosaccharide structures of permethylated glycans using a database library, FragLib, in conjunction with their Oligosaccharide Subtree Constraint Algorithm (OSCAR) [38,54,55]. This algorithm incorporates analyst-selected MS n ion fragmentation pathways for elucidation of oligosaccharide topology using a top-down sequencing strategy.…”

Discrimination of 16 structural isomers of fucosyl galactoside based on energy-resolved mass spectrometry

“…In most cases, there are differences in signal intensities, whereas in others, different fragment ions can be found. Accumulation of the spectral data in a database facilitates identification of oligosaccharides based on spectral matching [11][12][13][14][15][16][17]. An important problem-the elucidation of totally unknown structures that are not present in any databasewill still remain after the introduction of this efficient methodology.…”