Self-assembly of temperature-responsive di-block polypeptides functionalized with unnatural amino acids.

The incorporation of unnatural amino acids (uAAs) into protein-based polymers has emerged as a powerful methodology to expand their chemical repertoire. Recently, we demonstrated that incorporating uAAs into two temperature-responsive protein-based polymers-namely resilin- and elastin-like polypeptides (RLPs and ELPs, respectively)-can alter their properties. In this study, we incorporated aromatic uAAs into the protein sequence of RLP-ELP diblocks to yield new and diverse assemblies from a single DNA template.

Robust Photocontrol of Elastin-like Polypeptide Phase Transition with a Genetically Encoded Arylazopyrazole.

The rational design of light-responsive proteins and protein-based polymers requires both a photoswitch with suitable light-responsive properties and the ability to incorporate it at (multiple) defined positions in the protein chain. This Letter describes the evolution of high-performance aminoacyl-tRNA synthetases for recognizing a photoswitchable arylazopyrazole-bearing unnatural amino acid (AAP-uAA), which we then incorporated at multiple sites within elastin-like polypeptides (ELPs). The incorporation of AAP-uAA into ELPs yielded proteins capable of an isothermal, reversible, and robust light-mediated soluble-to-insoluble phase transition, which occurred faster (after only 1 min of light irradiation) and demonstrated a larger transition temperature difference (up to a 45 °C difference in the ELP transition temperature upon a cis to trans AAP isomerization) than similar azobenzene-containing ELPs.

Tuning the Properties of Protein-Based Polymers Using High-Performance Orthogonal Translation Systems for the Incorporation of Aromatic Non-Canonical Amino Acids.

The incorporation of non-canonical amino acids (ncAAs) using engineered aminoacyl-tRNA synthetases (aaRSs) has emerged as a powerful methodology to expand the chemical repertoire of proteins. However, the low efficiencies of typical aaRS variants limit the incorporation of ncAAs to only one or a few sites within a protein chain, hindering the design of protein-based polymers (PBPs) in which multi-site ncAA incorporation can be used to impart new properties and functions. Here, we determined the substrate specificities of 11 recently developed high-performance aaRS variants and identified those that enable an efficient multi-site incorporation of 15 different aromatic ncAAs.