Discovering functional, non-proteinogenic amino acid containing, peptides using genetic code reprogramming

Abstract

The protein synthesis machinery of the cell, the ribosome and associated factors, is able to accurately follow the canonical genetic code, that which maps RNA sequence to protein sequence, to assemble functional proteins from the twenty or so proteinogenic amino acids. A number of innovative methods have arisen to take advantage of this accurate, and efficient, machinery to direct the assembly of non-proteinogenic amino acids. We review and compare these routes to 'reprogram the genetic code' including in vitro translation, engineered aminoacyl tRNA synthetases, and RNA 'flexizymes'. These studies show that the ribosome is highly tolerant of unnatural amino acids, with hundreds of unusual substrates of varying structure and chemistries being incorporated into protein chains. We also discuss how these methods have been coupled to selection techniques, such as phage display and mRNA display, opening up an exciting new avenue for the production of proteins and peptides with properties and functions beyond that which is possible using proteins composed entirely of the proteinogenic amino acids.

Silicon amino acids

“…Although both experimental and theoretical investigations are mainly focused on the proteinogenic (natural) amino acids, the substantial relevance of unnatural amino acids has also been confirmed in the past. [2][3][4][5] These untypical compounds (e.g., D-amino acids, b-amino acids, as well as a-amino acids with modified side chains) are particularly used in the synthesis of modified peptides (such modifications result in the higher resistance toward biodegradation, the altered conformational flexibility, and changes in the higher-order structure of the peptide chains). [6,7] Due to their usefulness, the designing and synthesis of novel unnatural amino acids remain interesting research targets.…”

Rewriting the Genetic Code

“…The ribosome provides an intriguing target for accessing new genetically encoded polymers (30, 126). However, the key challenge that has prevented extensive ribosomal engineering has been the inaccessibility of orthogonal ribosomes.…”

“…These ncAAs are, however, often involved in cellular metabolism and posttranslational protein modification, as indicated in Figure 1 b . This review does not cover the promising genetic code expansion studies using in vitro protein synthesis (e.g., 126, 137), some of the established applications of in vivo genetic code engineering (15, 57, 77), or the exciting work with supernumerary unnatural base pairs (6, 14). …”

Max Bergmann award lecture:Macromolecular medicinal chemistry as applied to metabolic diseases

“…Decades later, ligation techniques have dramatically expanded the molecular range of synthetic targets from peptides to small proteins . Simultaneously, we have also witnessed site‐specific non‐native amino acid biosynthesis and ribosome‐free peptide synthesis providing molecular diversity that was historically restricted to small molecule synthetic chemistry.…”