The use of proteomic technologies to characterize and study the proteome of mycobacteria has provided important information in terms of function, diversity, protein-protein interactions, and host-pathogen interactions in Mycobacterium spp. There are many different mass spectrometry methodologies that can be applied to proteomics studies of mycobacteria and microorganisms in general. Sample processing and appropriate study design are critical to generating high-quality data regardless of the mass spectrometry method applied. Appropriate study design relies on statistical rigor and data curation using bioinformatics approaches that are widely applicable regardless of the organism or system studied. Sample processing, on the other hand, is often a niched process specific to the physiology of the organism or system under investigation. Therefore, in this chapter, we will provide protocols for processing mycobacterial protein samples for the specific application of Top-down and Bottom-up proteomic analyses.
Keywords: Bottom-Up Proteomics; ESI (Electro-Spray Ionization); LC-MS (Liquid Chromatography - Mass Spectrometry); MALDI-ToF (Matrix Assisted Laser Desorption Ionization - Time of Flight); MRM (Multiple Reaction Monitoring); PTM (Post-translational modification); Sample processing; Shotgun proteomics; Top-Down Proteomics.