The bacteria Helicobacter pylori is the leading cause of duodenal ulcers, gastritis, and gastric cancer worldwide. The sugars, or glycans, present on the surface of H. pylori are directly linked to this organism’s ability to be a pathogen; however, the structures of these sugars remain largely unknown. We are interested in determining glycan structures in order to aid the development of novel therapeutics to eradicate H. pylori, which is becoming increasingly resistant to current antibiotic treatments. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) is a powerful tool for structural identification; however, previous attempts to gather structural information about the glycans using this instrument have proven unsuccessful as the MS/MS spectra produced were dominated by peaks corresponding to the phosphine-based probes used to isolate the glycans. We hypothesized that the low signal intensity of the glycan peaks was due to a lack of basic sites on the sugars where a proton could be added, thus decreasing the LC-MS/MS’s sensitivity to the glycan. Therefore, reductive amination was employed to add basic sites onto the glycans. We found that reductively aminating synthesized glycan-phosphine-FLAG samples of GlcNAz-Phos-FLAG using N,N-dimethyl-p-phenylenediamine as the amine produced MS/MS spectra with abundant peaks corresponding to the reductively aminated GlcNAz portion of the ions. Based on the analysis of the product ions, we have proposed a fragmentation pathway for the dissociation of reductively aminated GlcNAz-Phos-FLAG, validating this method as a means by which structural information can be obtained from the glycans of Hp.

Addition of basic sites to the glycans of Helicobacter pylori to increase MS/MS peak abundance

251st National Meeting (2016)

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