Question

Explain the technique of peptide mapping used to compare normal haemoglobin with sickle cell haemoglobin.

Hint:

Peptide mapping identifies differences in protein structure by comparing enzyme-generated peptide fragments, useful for detecting mutations like in sickle cell haemoglobin.

Answer 1

Peptide Mapping is a biochemical technique used to analyze and compare proteins by breaking them down into smaller peptide fragments and studying their patterns. It helps detect structural differences caused by mutations.

• Both normal haemoglobin and sickle cell haemoglobin proteins are treated with a proteolytic enzyme such as trypsin which cleaves the protein at specific amino acid residues.
• This digestion produces smaller peptide fragments specific to the protein’s sequence.
• The peptide fragments are then separated by chromatography or electrophoresis based on their size and charge.
• The resulting peptide pattern (peptide map) is visualized, usually by staining or autoradiography.
• In sickle cell haemoglobin, a single amino acid substitution (glutamic acid replaced by valine) causes a change in the peptide fragments, which leads to a different pattern compared to normal haemoglobin.
• This difference in peptide mapping confirms the molecular basis of sickle cell anaemia.