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Efficient Phage Display with Multiple Distinct Non-Canonical Amino Acids Using Orthogonal Ribosome-Mediated Genetic Code Expansion.

ABSTRACT: Phage display is a powerful approach for evolving proteins and peptides with new functions, but the properties of the molecules that can be evolved are limited by the chemical diversity encoded. Herein, we report a system for incorporating non-canonical amino acids (ncAAs) into proteins displayed on phage using the pyrrolysyl-tRNA synthetase/tRNA pair. We improve the efficiency of ncAA incorporation using an evolved orthogonal ribosome (riboQ1), and encode a cyclopropene-containing ncAA (CypK) at diverse sites on a displayed single-chain antibody variable fragment (ScFv), in response to amber and quadruplet codons. CypK and an alkyne-containing ncAA are incorporated at distinct sites, enabling the double labeling of ScFv with distinct probes, through mutually orthogonal reactions, in a one-pot procedure. These advances expand the number of functionalities that can be encoded on phage-displayed proteins and provide a foundation to further expand the scope of phage display applications.

SUBMITTER: Oller-Salvia B 

PROVIDER: S-EPMC6771915 | biostudies-literature | 2019 Aug

REPOSITORIES: biostudies-literature

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Efficient Phage Display with Multiple Distinct Non-Canonical Amino Acids Using Orthogonal Ribosome-Mediated Genetic Code Expansion.

Oller-Salvia Benjamí B   Chin Jason W JW  

Angewandte Chemie (International ed. in English) 20190704 32

Phage display is a powerful approach for evolving proteins and peptides with new functions, but the properties of the molecules that can be evolved are limited by the chemical diversity encoded. Herein, we report a system for incorporating non-canonical amino acids (ncAAs) into proteins displayed on phage using the pyrrolysyl-tRNA synthetase/tRNA pair. We improve the efficiency of ncAA incorporation using an evolved orthogonal ribosome (riboQ1), and encode a cyclopropene-containing ncAA (CypK) a  ...[more]

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