Semi-Quantitative Analysis of Peptidoglycan by Liquid Chromatography Mass Spectrometry and Bioinformatics

Peptoglycomics uses advanced mass spectrometry combined with high-powered bioinformatic data analysis to examine peptidoglycan composition in fine detail. Using bioinformatic algorithms enables the analysis, extraction, and comparison of mass spec data across a collection of data files simultaneously, decreasing the workload and increasing the accuracy. Understanding the diverse variations and modifications of peptidoglycan has implications in understanding cellular processes, including growth and division, host immune evasion, and antimicrobial resistance.

We recommend preparing and analyzing a small subset of samples first to become familiar with the protocol and to act as a pilot project to assess the experimental parameters. As mass spectrometry data analysis software often varies in the definitions that are used when describing features and functions, visual demonstration will aid users in applying this protocol across platforms. For peptidoglycan sacculi extraction, resuspend frozen cell pellets at approximately one to 10 of the original culture volume in 20 millimolar sodium phosphate at four degrees Celsius before adding the cell suspension dropwise to boiling 8%SDS in 20 millimolar sodium phosphate to a final one-to-one volume.