Protein sequencing Edman reaction

The general idea of peptide sequencing by Edman degradation is to cleave one amino acid at a time from an end of the peptide chain. That terminal amino acid is then separated and identified, and the cleavage reactions are repeated on the chain-shortened peptide until the entire peptide sequence is known. Automated protein sequencers are available that allow as many as 50 repetitive sequencing cycles to be carried out before a buildup of unwanted by products interferes with the results. So efficient are these instruments that sequence information can be obtained from as little as 1 to 5 picomoles of sample—less than 0.1 /xg. 

THE EDMAN REACTION ENABLES PEPTIDES PROTEINS TO BE SEQUENCED... 

The unstable thiazolinones are converted into stable hydantoines in order to facilitate their identification. Conversion and identification are carried out outside the instrument after extraction of the thiazolinones with butylchlor-ide. The conversion reaction as well as the problems associated with identification of the PTH-amino acids were studied in detail by Edman and described explicitly in Needleman s book on Protein Sequence Determination Conversion is generally carried out in 1 N HCl at 80 °C within 10 min. The PTH-derivatives are extracted from the aqueous phase with ethyl acetate with the exception of PTH-arginine, PTH-histidine and PTH-cysteine which remain in the aqueous phase. 

B 2. The new reagent 4-W,W-dimethylaminoazobenzene-4 -isothiocyanate (DABITC) is often used for the Edman method of protein sequence analysis. Write a reaction showing this use of the reagent. 

Many proteins contain hundreds of amino acids. To determine their sequences other reactions are required to provide sequences short enough to be determined by Edman degradation. Enzymatic cleavage by two enzymes, trypsin and chymotrypsin, is used to produce smaller peptides. Trypsin cleaves polypeptide chains on the C-terminal side of basic residues such as arginine and lysine. Chymotrypsin cleaves the polypeptide on the C-terminal side of aromatic residues. 

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