Protein Digestion Techniques

Abstract

Modern MS instruments are capable of measuring the molecular weights of intact proteins with a fairly high degree of accuracy. So why not do proteomics simply by measuring the masses of intact proteins? Unfortunately, intact mass measurements are of relatively little use for three reasons. First, as good as MS instruments are, there are still errors in the measurements they produce. The greater the mass of the protein, the greater the absolute magnitude of the error. Those errors introduce enough uncertainty to make the measurements insufficiently accurate for definitive identification. Moreover, diverse posttranslational modifications further complicate assignments based on mass. Second, not all proteins are amenable to intact mass measurements. It can be very difficult to obtain mass measurements on very large and hydrophobic proteins. Third, the sensitivity of measurements of intact protein masses is not nearly as good as sensitivity for peptide mass measurements and peptide tandem MS analyses. For these reasons, doing proteomics by analyzing intact proteins is not a realistic option at present.