Bottom-up proteomics

Proteomics

Proteomics is the large-scale study of proteins, particularly their structures and functions. Proteins are vital parts of living organisms, as they are the main components of the physiological metabolic pathways of cells. The term "proteomics" was first coined in 1997 to make an analogy with...

is a common method to identify proteins and characterize their amino acid sequences and post-translational modifications by proteolytic digestion

Proteolysis

Proteolysis is the directed degradation of proteins by cellular enzymes called proteases or by intramolecular digestion.-Purposes:Proteolysis is used by the cell for several purposes...

 of proteins prior to analysis by mass spectrometry

Mass spectrometry

Mass spectrometry is an analytical technique that measures the mass-to-charge ratio of charged particles.It is used for determining masses of particles, for determining the elemental composition of a sample or molecule, and for elucidating the chemical structures of molecules, such as peptides and...

. The proteins may first be purified by a method such as gel electrophoresis

Gel electrophoresis

Gel electrophoresis is a method used in clinical chemistry to separate proteins by charge and or size and in biochemistry and molecular biology to separate a mixed population of DNA and RNA fragments by length, to estimate the size of DNA and RNA fragments or to separate proteins by charge...

 resulting in one or a few proteins in each proteolytic digest. Alternatively, the crude protein extract is digested directly, followed by one or more dimensions of separation of the peptides by liquid chromatography

High performance liquid chromatography

High-performance liquid chromatography , HPLC, is a chromatographic technique that can separate a mixture of compounds and is used in biochemistry and analytical chemistry to identify, quantify and purify the individual components of the mixture.HPLC typically utilizes different types of stationary...

 coupled to mass spectrometry, a technique known as shotgun proteomics

Shotgun proteomics

Shotgun proteomics is a method of identifying proteins in complex mixtures using a combination of high performance liquid chromatography combined with mass spectrometry. The name is derived from shotgun sequencing of DNA which is itself named by analogy with the rapidly-expanding, quasi-random...

. By comparing the masses of the proteolytic peptides or their tandem mass spectra

Tandem mass spectrometry

Tandem mass spectrometry, also known as MS/MS or MS2, involves multiple steps of mass spectrometry selection, with some form of fragmentation occurring in between the stages.-Tandem MS instruments:...

 with those predicted from a sequence database or annotated peptide spectral in a peptide spectral library

Peptide Spectral Library

Peptide Spectral Library is a curated, annotated and non-redundant collection/database of LC-MS/MS peptide spectra. One essential utility of Peptide Spectral Library is to serve as consensus templates supporting the identification of peptide/proteins based on the correlation between the templates...

, peptides can be identified and multiple peptide identifications assembled into a protein identification.