N-Terminal Amino Acid Sequencing and Edman Degradation Overview

Amino acid N-terminal sequencing is a technique used to determine the order of several amino acid residues at the start of a protein or polypeptide sequence from the N-terminus (i.e., the amino terminal). This method is extremely important in protein identification and characteristic analysis, especially in biochemical and molecular biological research.

 

The Edman degradation method is the most commonly used amino acid N-terminal sequencing method. This method determines the amino acid sequence of a protein or polypeptide by chemically degrading the sequence one amino acid at a time. The process is roughly as follows.

 

Labeling N-Terminal Amino Acids

First, a specific chemical reagent (usually Edman's reagent, phenylisothiocyanate) reacts with the N-terminal amino acid of the polypeptide or protein. This reaction forms a stable labeled compound.

 

Removal of Labeled Amino Acids

Next, a special chemical method is used to remove this labeled amino acid from the polypeptide chain. The removed amino acid is a stable derivative that can be analyzed and identified by techniques such as high-performance liquid chromatography (HPLC).

 

Repeat the Process

Then the process is repeated with the remaining polypeptide or protein, removing a new N-terminal amino acid evrytime. By repeating this process step by step, the entire sequence of the polypeptide or protein can be determined.

 

It should be noted that the efficiency of Edman degradation is not 100%, which means that the signal will gradually weaken as the sequencing depth increases. Therefore, this method is typically only used to determine the first 20-30 amino acids of a protein. For long-chain proteins or situations where the entire sequence needs to be determined, other methods, such as mass spectrometry, are usually combined.