When can glycopeptides be assigned based solely on high-resolution mass spectrometry data?

Glycoproteomics is an emerging science that shows promise in applications such as biomarker discovery and biopharmaceutical development. One central technique in glycoproteomic analysis is analyzing glycopeptides by mass spectrometry. This challenging technique is still under development, and methods to simplify the data analysis are greatly needed. One potentially attractive analysis approach would be to assign a significant portion of the glycopeptide compositions using high-resolution MS data. In the work described herein, we ask the question: Under what circumstances is it possible to assign glycopeptides to MS data, using only high-resolution mass spectra? Variables investigated include the number of glycosylation sites on the protein, the potential diversity of the glycans attached to the protein, and the mass accuracy obtained. This work outlines guidelines for when it is (and is not) appropriate to rely heavily on high-resolution mass measurements to assign glycopeptide compositions; such guidelines are potentially useful for anyone conducting glycopeptide analysis by mass spectrometry.