Solid Phase Polypeptide Synthesis
Merrifield Solid Phase Polypeptide Synthesis
The Merrifield solid-phase peptide synthesis (SPPS) technique was developed by R. Bruce Merrifield in the 1960s. The Merrifield method involves the use of a solid support, which consists of beads of a copolymer. The copolymer is synthesized from reaction of styrene with 4-(chloronmethyl) styrene.
The synthesis involves in the following steps:
Step 1. The N-terminal protected (BoC) amino acid has been attached to the copolymer through ester bond formation between -COOH of amino acid and -CH2Cl of the copolymer.
Step 2. The copolymer attached to amino acid is washed to remove excess reagents. The reaction with trifluoroacetic acid (CH3COOH) is then carried out to remove BoC protecting group so that a free amino group becomes available.
Step 3. Another amino acid (amino group protected with BoC) is then added to copolymer in the presence of DCC. The free amino group of amino acid attached to polymer forms peptide linkage with free carboxylic group of second amino acid.
Step 4. Copolymer and the attached dipeptide is again washed for removing excess reagent for the addition of another amino acid. Steps 2 and 3 are now repeated. These steps can be repeated a number of times depending upon the length of polypeptide chain.
Step 5. Once the required polypeptide is synthesized by linking of different amino acids the polypeptide is released from the copolymer by reaction with hydrofluoric acid (HF). Hydrofluoric acid is having two functions:
(a) It cleaves the ester bond and leaves the peptide chain intact.
(b) It removes the N-terminal protecting group, namely BoC.
The Merrifield solid phase peptide synthesis is presently carried out with automated peptide synthesizer which automatically undertakes the washing and removal of protecting group at each addition step.
Source: Biochemistry by C.B. POWAR and G.R. CHATWAL
