How to interpret the mass spectrogram of peptides
Mass spectrometry of peptides is an important tool for identifying proteins and peptide fragments, but correctly understanding and interpreting these images may require some basic knowledge and experience. Interpreting peptide mass spectrometry generally involves six steps: understanding the mass spectrum, confirming ion types, sequencing, determining the order of amino acids, recognizing modifications, and verifying results.
1. Understanding the Mass Spectrum
A mass spectrum is a graphical representation of the relative abundance of ions with different masses in a sample. On the x-axis, we have the mass-to-charge ratio (m/z), and on the y-axis, we have relative abundance.
2. Confirming Ion Types
Peptide mass spectrometry typically shows b ions and y ions. B ions are generated from the N-terminal of the peptide, while y ions are generated from the C-terminal. These two types of ions dominate in peptide mass spectrometry.
3. Sequencing
By calculating the difference in m/z, we can determine the amino acid sequence of the peptide. For example, if the difference between two adjacent peaks corresponds to glycine (Gly), this indicates a glycine residue at that position.
4. Determining the Order of Amino Acids
By comparing the m/z differences of various peaks on the mass spectrum, we can determine the sequence of amino acids in the peptide. Typically, we start from lower mass peaks and move upwards; this is because lower mass peaks usually correspond to amino acids closer to the N-terminal of the peptide.
5. Recognizing Modifications
Some peptides may contain modifications such as phosphorylation or acetylation, which typically increase the mass of the peptide and produce characteristic peaks on the mass spectrum. If the m/z difference does not match the mass of any amino acid, it may be due to modifications.
6. Verifying Results
Finally, you can use software or online tools, such as PeptideMass, to verify your interpretation results. These tools can calculate the theoretical m/z values of your peptide sequence, which you can compare with experimental results.
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