Utility of Ion Mobility – Mass Spectrometry in Peptidoglycan Structural Determination

The peptidoglycan (PG) found within bacterial cell walls is a complex megastructure with components that are difficult to identify without significant time commitment. Potential isomers, modifications, and cross-linking patterns within whole cell digests lead to complex samples that typically require manual analysis. The introduction of a method for streamlining the identification of peptidoglycan components (muropeptides) is needed to develop a better structure identification workflow. This work explores trapped ion mobility – mass spectrometry as a tool to provide muropeptide structural information through analysis of ion mobilities and collision cross section (CCS). PG of two filamentous cyanobacteria, Nostoc commune and Limnospira platensis, were chosen for this work as they have not been characterized before now despite commonplace use as sources of nutrition. The analyses revealed that muropeptides can be organized into distinct regions of similar cross-linking structure by comparing their CCS and m/z values. Both species possess typical Gram-negative structures in-line with their classifications as Gram-negative species. The PG of L. platensis was found to contain agmatine as a modification to the D-iso-glutamic acid residue within the peptide stem. This modification was found to exist within most (~90 %) of its identified muropeptides, indicating that agmatine may play an important role in L. platensis growth and development.