Protein sequencing determines the order of amino acids in a protein. Among chemical methods N-terminal sequencing by Edman degradation is by far the most commonly used. By this method we get information about the order of amino acids from the amino-terminal end.
Sequence information can be used to identify a protein or homologous proteins through searches in databases; the minimal number of residues for a succesful search is ten, but often more residues are required. The N-terminal sequence is often used for confirmation of the identity of a protein; in that case a few residues are enough. The sequence also gives information about post translational cleavage points. In addition, the sequence results give informations about the purity of a preparation; limits of detectable contamination depend on the sequences of the analysed proteins.
The most important prerequisite for a protein to be sequenced by Edman degradation is that the N-terminal residue is not blocked, but free to react in the first step of the sequencing procedure. Proteins can be blocked either naturally e.g. by acetylation, or during purification by cyclisation of glutamine. It is estimated that >50% of all proteins are blocked. In these cases no sequence is obtained.
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Technician Anne Blicher (daily at 9-15)
Associate Professor Susanne Jacobsen
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