What is the Basic Principle of Identifying Proteins by Peptide Mass Mapping

What is the basic principle of identifying proteins by peptide mass mapping? The basic principle of peptide mass mapping for protein identification is the matching of many masses from different peptide fragments of a protein.

Peptide mass mapping is a technique that uses unique characteristics of proteins for identification. Therefore, it is also known as the peptide fingerprint.

Key Areas Covered

1. What is a Protein
     – Definition, Features, Importance
2. What is the Basic Principle of Identifying Proteins by Peptide Mass Mapping
     – Explanation of the Importance

Key Terms

Protein, Protein Mass Mapping

What is the Basic Principle of Identifying Proteins by Peptide Mass Mapping

What are Proteins

A protein is a large nitrogenous organic compound, composed of one or two amino acid chains. Hence, the building blocks of a protein are amino acids. A protein is made up of an alternative assembly of universal amino acids. In addition, peptide bonds are formed between amino groups and carboxyl acid groups of adjacent amino acids, forming a sequence of amino acids. Therefore, proteins are called polypeptides. This means protein is a polymer. Typically, a polypeptide consists of 50 – 2000 amino acids.

What is the Basic Principle of Identifying Proteins by Peptide Mass Mapping

Figure 1: Protein Structure

Furthermore, protein synthesis occurs as a result of gene expression. Transcription and translation are the two steps of protein synthesis. The study of the structure and function of these proteins is called proteomics. Proteins are a very complex and dynamic type of molecule. A typical protein consists of four structural levels: primary, secondary, tertiary, and quaternary. Different combinations of amino acids give different properties to proteins. Humans have 20,000 to 25,000 protein-coding genes. About 2 million different protein types can be synthesized from them. The human body contains around 50,000 proteins. The rest of the proteins are consumed throughout the diet.

What is the Basic Principle of Identifying Proteins by Peptide Mass Mapping

Peptide mass mapping is an analytical technique for protein identification. It uses powerful search engines in order to identify proteins from mass spectrometry data. The basic principle of peptide mass mapping is the digestion of a small portion of a protein by a proteolytic enzyme such as trypsin. Here, trypsin cleaves proteins at lysine and arginine residues. Importantly, 10% of a protein is composed of lysine and arginine residues in a typical protein. Therefore, it results in peptide fragments of 500 to 3000 Da in size. Mass spectrometry analyzes the mixture resulting from proteinase cleavage. Then, it compares the experimental and theoretical mass spectrometry values. The theoretical mass spectrometry values are taken by genome sequencing. It allows the prediction of all proteins in the genome.

Identifying Proteins by Peptide Mass Mapping

Figure 2; Peptide Mass Mapping

Further, depending on the cleavage rules for the digestion of proteins, mass values of the peptides from cleavage are determined. An appropriate scoring algorithm then compares the masses of experimental peptides with the theoretical peptide mass values of proteins. This allows the identification of proteins. Apart from that, unknown proteins can match a particular protein in the database. Homology is applicable to identify unknown proteins with the use of peptide databases.

Meanwhile, the analysis of a complex protein mixture always uses a separation step to isolate a certain protein. Two-dimensional gel electrophoresis or liquid chromatography involves in this step.


In brief, proteins are large, organic compounds composed of carbon, nitrogen, oxygen, hydrogen, and sulfur-like atoms. A polypeptide chain contains around 50 to 2000 amino acids. Peptide mass mapping is the analytical technique responsible for the identification of proteins depending on the peptide fingerprint of the protein. Therefore, it involves the cleavage of protein by a proteolytic enzyme like trypsin. The mass spectrometry analyzes the resulting peptides. Subsequently, it compares the experimental peptide masses with the theoretical peptide masses of proteins. This method identifies the exact protein or proteins based on homology.

  1. Thiede B, Höhenwarter W, Krah A, Mattow J, Schmid M, Schmidt F, Jungblut PR. Peptide mass fingerprinting. Methods. 2005 Mar;35(3):237-47. doi: 10.1016/j.ymeth.2004.08.015. Epub 2005 Jan 12. PMID: 15722220.
Image Courtesy:
  1. Main protein structure levels en” By LadyofHats– Derivative work (Public Domain) via Commons Wikimedia
  2. Peptide mass fig” By Lfont39 – Own Work (CC-BY-SA 4.0) via Commons Wikimedia

About the Author: Lakna

Lakna, a graduate in Molecular Biology & Biochemistry, is a Molecular Biologist and has a broad and keen interest in the discovery of nature related things

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