TY - JOUR

T1 - Nonproteinogenic deep mutational scanning of linear and cyclic peptides

AU - Rogers, Joseph M.

AU - Passioura, Toby

AU - Suga, Hiroaki

PY - 2018/10/23

Y1 - 2018/10/23

N2 - High-resolution structure-activity analysis of polypeptides requires amino acid structures that are not present in the universal genetic code. Examination of peptide and protein interactions with this resolution has been limited by the need to individually synthesize and test peptides containing nonproteinogenic amino acids. We describe a method to scan entire peptide sequences with multiple nonproteinogenic amino acids and, in parallel, determine the thermodynamics of binding to a partner protein. By coupling genetic code reprogramming to deep mutational scanning, any number of amino acids can be exhaustively substituted into peptides, and single experiments can return all free energy changes of binding. We validate this approach by scanning two model protein-binding peptides with 21 diverse nonproteinogenic amino acids. Dense structure-activity maps were produced at the resolution of single aliphatic atom insertions and deletions. This permits rapid interrogation of interaction interfaces, as well as optimization of affinity, fine-tuning of physical properties, and systematic assessment of nonproteinogenic amino acids in binding and folding.

AB - High-resolution structure-activity analysis of polypeptides requires amino acid structures that are not present in the universal genetic code. Examination of peptide and protein interactions with this resolution has been limited by the need to individually synthesize and test peptides containing nonproteinogenic amino acids. We describe a method to scan entire peptide sequences with multiple nonproteinogenic amino acids and, in parallel, determine the thermodynamics of binding to a partner protein. By coupling genetic code reprogramming to deep mutational scanning, any number of amino acids can be exhaustively substituted into peptides, and single experiments can return all free energy changes of binding. We validate this approach by scanning two model protein-binding peptides with 21 diverse nonproteinogenic amino acids. Dense structure-activity maps were produced at the resolution of single aliphatic atom insertions and deletions. This permits rapid interrogation of interaction interfaces, as well as optimization of affinity, fine-tuning of physical properties, and systematic assessment of nonproteinogenic amino acids in binding and folding.

KW - BH3 domains

KW - Intrinsically disordered proteins

KW - Macrocyclic peptides

KW - Noncanonical amino acids

KW - Structure-activity relationships

UR - http://www.scopus.com/inward/record.url?scp=85055524065&partnerID=8YFLogxK

U2 - 10.1073/pnas.1809901115

DO - 10.1073/pnas.1809901115

M3 - Journal article

C2 - 30301798

AN - SCOPUS:85055524065

VL - 115

SP - 10959

EP - 10964

JO - Proceedings of the National Academy of Sciences of the United States of America

JF - Proceedings of the National Academy of Sciences of the United States of America

SN - 0027-8424

IS - 43

ER -