Optimizing SureQuant for Targeted Peptide Quantification: a Technical Comparison with PRM and SWATH-MS Methods

Bacterial infections are a major threat to human health
worldwide.
A better understanding of the properties and physiology of bacterial
pathogens in human tissues is required to develop urgently needed
novel control strategies. Mass spectrometry-based proteomics could
yield such data, but identifying and quantifying scarce bacterial
proteins against a preponderance of human proteins is challenging.
Here, we explored the recently introduced SureQuant method for highly
sensitive targeted mass spectrometry. Using a major human pathogen,
the Gram-positive bacteria Staphylococcus aureus, as an example, we evaluated several parameters, including the number
of targets and intensity thresholds, for optimal qualitative and quantitative
protein analysis. By comparison, we found that SureQuant achieved
the same quantitative performance as standard parallel reaction monitoring
while allowing accurate and precise quantification of up to 400 targets.
SureQuant also surpassed the sensitivity and quantification capabilities
of global data-independent acquisition methods. Finally, to facilitate
method development, we provide optimized MS parameters for the sensitive
quantification of different peptide panel sizes. This study provides
a foundation for the broader application of SureQuant in the analysis
of clinical specimens containing trace amounts of bacterial proteins
as well as other studies requiring ultrasensitive detection of low-abundant
proteins.