Peptide Synthesis
A peptide is composed of two or more amino acids linked by an amide bond, forming a chain typically ranging from 2 to 70 amino acids in length. Unlike proteins, peptides do not require folding to be biologically active. Peptides are naturally occurring as peptide hormones such as angiotensin, LHRH, and enkephalin, and as toxins in plants and animals. They are of significant interest in drug discovery as potential lead compounds, and also serve as drugs themselves. In addition, peptides are widely used in vaccines, biomaterials, histological probes, and as antigens to generate antibodies. Peptides can be synthesized chemically either in solution or via solid-phase synthesis, where the process involves the selective formation of amide bonds between protected amino acids. In solid-phase synthesis, the carboxyl group is protected and linked to a polymer support. After bond formation, the amino-protecting group of the dipeptide is removed, and the next N-protected amino acid is coupled to the growing chain.
Overview
Solid-Phase Peptide Synthesis (SPPS)
Solid-phase peptide synthesis (SPPS) is the most frequently used method of peptide synthesis due to its efficiency, simplicity, speed, and ease of parallelization. SPPS involves sequential addition of amino and side-chain protected amino acid residues to an amino acid or peptide attached to an insoluble polymeric support.
Boc and Fmoc Group Methods
Either an acid-labile Boc group (Boc SPPS) or base-labile Fmoc-group (Fmoc SPPS) is used for N-α-protection. After removal of this protecting group, the next protected amino acid is added using either a coupling reagent or pre-activated protected amino acid derivative.
C-terminal Anchoring and Side-Chain Protection
The C-terminal amino acid is anchored to the resin via a linker, the nature of which determines the conditions required to release the peptide from the support after chain extension. Side-chain protecting groups are often chosen so as to be cleaved simultaneously with detachment of the peptide from the resin.
Side-Chain Protecting Groups for SPPS
Side-chain protecting groups for solid-phase peptide synthesis (SPPS) are often chosen so as to be cleaved simultaneously with detachment of the peptide from the resin. Most peptides are prepared by the Fmoc method as the final cleavage and deprotection is carried out with trifluoroacetic acid.
Peptide Synthesis Limitations and Advances
Peptides of 50 amino acids can be routinely prepared, although the synthesis of proteins with over 100 amino acids is commonly reported. Longer proteins can be made by native chemical ligation of fully deprotected peptides in solution, enabling the synthesis of natural peptides, unnatural or D-amino acids, and peptides with cyclic, branched, labelled, and post-translational modifications.
Liquid-Phase Peptide Synthesis vs. Solid-Phase
Liquid-phase peptide synthesis, usually utilizing Boc or Z-amino protection, has been largely superseded by solid-phase peptide synthesis, except for existing processes of large-scale peptide synthesis for industrial purposes.