Edman degradation: Difference between revisions

Latest revision as of 11:22, 15 February 2025

Edman Degradation[edit]

Diagram of the Edman degradation process

The Edman degradation is a method of protein sequencing that allows for the sequential identification of amino acids in a peptide. This technique was developed by Pehr Edman in the 1950s and has been a fundamental tool in biochemistry for determining the primary structure of proteins.

Principle[edit]

The Edman degradation involves the selective cleavage of the N-terminal amino acid of a peptide, which is then identified as a phenylthiohydantoin (PTH) derivative. This process can be repeated multiple times to sequence the entire peptide.

Step 1: Coupling[edit]

In the first step, the peptide is treated with phenyl isothiocyanate (PITC) under mildly alkaline conditions. This reagent reacts with the free amino group of the N-terminal amino acid to form a phenylthiocarbamoyl derivative.

Step 2: Cleavage[edit]

The peptide is then treated with anhydrous acid, typically trifluoroacetic acid (TFA), which cleaves the N-terminal amino acid as a cyclic phenylthiohydantoin (PTH) derivative, leaving the rest of the peptide intact.

Step 3: Identification[edit]

The PTH-amino acid is identified using chromatography or mass spectrometry. This identification step is crucial for determining the sequence of the peptide.

Advantages and Limitations[edit]

The Edman degradation is advantageous for its ability to sequence peptides with high accuracy. However, it has limitations, such as the inability to sequence peptides longer than about 50 residues due to incomplete cleavage and side reactions. Additionally, it requires a free N-terminal amino group, which may not be available in all peptides.

Applications[edit]

Edman degradation has been widely used in the field of proteomics for the analysis of protein structure and function. It has been instrumental in the sequencing of many important proteins and in the study of enzyme mechanisms.

Related pages[edit]

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-'''Edman degradation''' is a method of sequencing [[amino acids]] in a [[peptide]]. It was developed by [[Pehr Edman]], hence the name. The method labels and removes only the amino-terminal (N-terminal) [[amino acid]] of the protein, which allows the identification of the amino acid.
+== Edman Degradation ==
-== Process ==
+[[File:EdmanDegradation.png|thumb|right|Diagram of the Edman degradation process]]
-The process of Edman degradation begins with the reaction of the N-terminal amino acid of the peptide with phenyl isothiocyanate under slightly alkaline conditions. This results in a phenylthiocarbamyl derivative. The peptide is then treated with anhydrous acid, which leads to the selective cleavage of the N-terminal amino acid as anilinothiazolinone derivative. The derivative can be easily identified by chromatography or electrophoresis. The remaining peptide, now shorter by one amino acid, can then undergo another round of degradation.
+The '''Edman degradation''' is a method of [[protein]] sequencing that allows for the sequential identification of [[amino acids]] in a [[peptide]]. This technique was developed by [[Pehr Edman]] in the 1950s and has been a fundamental tool in [[biochemistry]] for determining the [[primary structure]] of proteins.
-== Limitations ==
+== Principle ==
-While Edman degradation is a powerful tool for determining the N-terminal sequence of a peptide, it has several limitations. The method is relatively slow, with each cycle taking around 1 hour. It also requires a large amount of the peptide to be effective. Furthermore, the method can only reliably sequence up to 50 amino acids. For larger peptides, other methods such as [[mass spectrometry]] are more effective.
+The Edman degradation involves the selective cleavage of the [[N-terminal]] amino acid of a peptide, which is then identified as a [[phenylthiohydantoin]] (PTH) derivative. This process can be repeated multiple times to sequence the entire peptide.
+=== Step 1: Coupling ===
+In the first step, the peptide is treated with [[phenyl isothiocyanate]] (PITC) under mildly alkaline conditions. This reagent reacts with the free amino group of the N-terminal amino acid to form a phenylthiocarbamoyl derivative.
+=== Step 2: Cleavage ===
+The peptide is then treated with anhydrous [[acid]], typically [[trifluoroacetic acid]] (TFA), which cleaves the N-terminal amino acid as a cyclic [[phenylthiohydantoin]] (PTH) derivative, leaving the rest of the peptide intact.
+=== Step 3: Identification ===
+The PTH-amino acid is identified using [[chromatography]] or [[mass spectrometry]]. This identification step is crucial for determining the sequence of the peptide.
+== Advantages and Limitations ==
+The Edman degradation is advantageous for its ability to sequence peptides with high accuracy. However, it has limitations, such as the inability to sequence peptides longer than about 50 residues due to incomplete cleavage and side reactions. Additionally, it requires a free N-terminal amino group, which may not be available in all peptides.
 == Applications ==
-Despite its limitations, Edman degradation has found wide use in the field of [[biochemistry]]. It is commonly used to determine the primary structure of small proteins. It can also be used to identify proteins or to determine the boundaries of a protein domain.
+Edman degradation has been widely used in the field of [[proteomics]] for the analysis of [[protein structure]] and function. It has been instrumental in the sequencing of many important proteins and in the study of [[enzyme]] mechanisms.
+== Related pages ==
-== See also ==
 * [[Protein sequencing]]
-* [[Mass spectrometry]]
+* [[Amino acid]]
-* [[Pehr Edman]]
+* [[Peptide]]
+* [[Phenyl isothiocyanate]]
+* [[Chromatography]]
-[[Category:Biochemistry methods]]
+[[Category:Biochemistry]]
-[[Category:Protein sequencing]]
+[[Category:Protein methods]]
-{{biochemistry-stub}}