Bottom-up proteomics

Bottom-up proteomics is a common method to identify proteins and characterize their amino acid sequences and post-translational modifications by proteolytic digestion of proteins prior to analysis by mass spectrometry.^[1][2] BUP techniques can be an alternative to MALDI-TOF MS approaches, as they allow the identification of bacterial strains and the characterization of potential resistance and virulence factors in a single run.^[3] The major alternative workflow used in high-throughput proteomics is called top-down proteomics and does not use proteolytic digestion. Essentially, bottom-up proteomics is a relatively simple and reliable means of determining the protein make-up of a given sample of cells, tissues, etc.^[4]

In bottom-up proteomics, the crude protein extract is enzymatically digested, followed by one or more dimensions of separation of the peptides by liquid chromatography coupled to mass spectrometry, a technique known as shotgun proteomics.^[5][6] By comparing the masses of the proteolytic peptides or their tandem mass spectra with those predicted from a sequence database or annotated peptide spectral in a peptide spectral library, peptides can be identified and multiple peptide identifications assembled into a protein identification.