Peptide Mapping

Abstract

Proteomics is the large‐scale study of proteins, particularly their structures and functions. One of its most important tasks is to develop efficient and rapid approaches to identify various proteins. Peptide mapping is a commonly used strategy in protein identification. It involves the chemical or enzymatic treatment of proteins resulting in the formation of peptide fragments followed by separation and identification of the resultant fragments. Proteins are usually digested into peptides that are subsequently identified by mass spectrometry to obtain peptide mass fingerprints. Peptide mapping is an important technique for investigating protein primary structures and determining surface‐exposed sites or epitopes within proteins. It can be adapted to obtain internal protein sequences. Prior to identification, the proteins and peptides usually need to be separated by a variety of separation approaches, such as gel electrophoresis, liquid chromatograph, capillary electrophoresis, etc.

Key Concepts:

  • Sodium dodecyl sulphate–polyacrylamide gel electrophoresis (SDS‐PAGE) is a commonly used approach for the separation of proteins and peptides.

  • Two‐dimensional gel electrophoresis (2‐DE) can separate a great many of proteins in two separation modes.

  • Proteolysis is a process where proteins are broken down into smaller peptides by proteases.

  • Proteases are enzymes that conduct proteolysis by hydrolysis of some specific peptide bonds.

  • Mass spectroscopy (MS) is an analytical technique to separate the components of a sample by their mass‐to‐charge ratios.

  • Peptide mass fingerprinting is the most popular analytical application of matrix‐assisted laser desorption/ionisation‐time of flight MS (MALDI‐TOF MS) and electrospray MS.

  • Electrospray became widely used as ionisation source for the MS analysis of proteins and peptides.

  • Mascot is a powerful search engine which uses mass spectrometry data to identify proteins from primary sequence databases.

  • Sequence coverage is the percentage of the database protein sequence covered by matching peptides.

  • Proteolytic microchip bioreactors are microdevices in which proteases are immobilised for efficient peptide mapping.

Keywords: peptide; separation; SDS‐PAGE; 2‐DE; mass spectrometry; peptide mass fingerprinting

Figure 1.

Schematic diagrams showing (a) a typical microchip proteolysis system, (b) the amplified channel in a microchip bioreactor and (c) the photograph of a microchip bioreactor.

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References

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Further Reading

Bao HM, Chen QW, Zhang LY and Chen G (2011) Immobilization of trypsin in the layer‐by‐layer coating of graphene oxide and chitosan on in‐channel glass fiber for microfluidic proteolysis. Analyst 136: 5190–5196.

Bao HM, Lui T, Zhang LY and Chen G (2009) Infrared‐assisted proteolysis using trypsin‐immobilized silica microspheres for peptide mapping. Proteomics 9: 1114–1117.

Fan HZ, Bao HM, Zhang LY and Chen G (2011) Immobilization of trypsin on poly(urea‐formaldehyde)‐coated fiberglass cores in microchip for highly efficient proteolysis. Proteomics 11: 3420–3423.

Fan HZ, Chen Z, Zhang LY, Yang PY and Chen G (2008) Fabrication and performance of poly(methyl methacrylate) microfluidic chips with fiber cores. Journal of Chromatography A 1179: 224–228.

Lauber MA, Koza SM, McCall SA et al. (2013) High‐resolution peptide mapping separations with MS‐friendly mobile phases and charge‐surface‐modified C18. Analytical Chemistry 85: 6936–6944.

Liu T, Bao HM and Chen G (2010) Inflation bulb‐driven microfluidic reactor for infrared‐assisted proteolysis. Electrophoresis 31: 3070–3073.

Liu T, Bao HM, Zhang LY and Chen G (2009) Integration of electrodes in a suction cup‐driven microchip for alternating current‐accelerated proteolysis. Electrophoresis 30: 3265–3268.

Liu T, Wang S and Chen G (2009) Immobilization of trypsin on silica‐coated fiberglass core in microchip for highly efficient proteolysis. Talanta 77: 1767–1773.

Wang S, Bao HM, Yang PY and Chen G (2008) Infrared‐assisted on‐plate proteolysis for MALDI‐TOF‐MS peptide mapping. Analytical Chemistry 80: 5640–5647.

Wang S, Chen Z, Yang PY and Chen G (2008) Trypsin‐immobilized fiber core in syringe needle for highly efficient proteolysis. Proteomics 8: 1785–1788.

Wang S, Liu T, Zhang LY and Chen G (2009) Pengyuan Yang, Chymotryptic proteolysis accelerated by alternating current for MALDI‐TOF‐MS peptide mapping. Journal of Proteomics 72: 640–647.

Whitmore CD and Gennaro LA (2012) Capillary electrophoresis‐mass spectrometry methods for tryptic peptide mapping of therapeutic antibodies. Electrophoresis 33: 1550–1556.

Xie H, Gilar M and Gebler JC (2009) Characterization of protein impurities and site‐specific modifications using peptide mapping with liquid chromatography and data independent acquisition mass spectrometry. Analytical Chemistry 81: 5699–5708.

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How to Cite close
Chen, Gang(Apr 2014) Peptide Mapping. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1002/9780470015902.a0002675.pub2]