Expanding the chemical toolbox for the synthesis of large and uniquely modified proteins

Abstract

Methods to prepare proteins that include a specific modification at a desired position are essential for understanding their cellular functions and physical properties in living systems. Chemical protein synthesis, which relies on the chemoselective ligation of unprotected peptides, enables the preparation of modified proteins that are not easily fabricated by other methods. In contrast to recombinant approaches, chemical synthesis can be used to prepare protein analogues such as D-proteins, which are useful in protein structure determination and the discovery of novel therapeutics. Post-translationally modifying proteins is another example where chemical protein synthesis proved itself as a powerful approach for preparing samples with high homogeneity and in workable quantities. In this Review, we discuss the basic principles of the field, focusing on novel chemoselective peptide ligation approaches such as native chemical ligation and the recent advances based on this method with a proven record of success in the synthesis of highly important protein targets.

Accessing human selenoproteins through chemical protein synthesis

“…19–26 CPS is based mainly on solid-phase peptide synthesis (SPPS) 27 and chemoselective ligation reactions, 28–32 for example native chemical ligation (NCL). 28 Using this technology, it is possible to prepare (seleno)proteins of up to ∼200 amino acids.…”

Site-Specific Incorporation of a Thioester Containing Amino Acid into Proteins

“…1–2 To this end, a number of methods have been developed including the selective modification of cysteine residues with electrophiles, 3–4 the fusion of peptide tags to proteins of interest which can be subsequently chemically or enzymatically modified, 5–8 and the incorporation of noncanonical amino acids (ncAAs) into proteins through semisynthetic 9–11 or recombinant methods. 12–13 In nature, the thioester group provides a means for selective protein conjugation; examples include protein ubiquitination, 14 intein splicing, 15 and the attachment of complement factors to pathogens.…”

“…16 In protein chemistry, C-terminal thioesters have been very useful for the semisynthesis of protein variants by native chemical ligation. 11 While C-terminal protein thioesters may be accessed using recombinant intein-based technology, 15 the selective incorporation of thioesters at internal sites in recombinant proteins is difficult. Robust methods to do so would enable the formation of a variety of small molecule and polypeptide conjugates, including branched and lariat proteins.…”

Piperidine-based glycodendrons as protein N-glycan prosthetics

“…11,12 Indeed, modern innovations in chemical methods have made the total synthesis of functional glycoproteins a reality. 13 However, the synthetic methodology leading to glycan-modified polypeptides remains a challenging hurdle. To circumvent this issue, a hybrid approach has emerged in which recombinant proteins are produced in cell hosts and later chemically glycosylated.…”