Chemoenzymatic Posttranslational Modification Reactions for the Synthesis of Ψ[CH<sub>2</sub>NH]‐Containing Peptides

Kato, Yasuharu; Kuroda, Tomohiro; Huang, Yichao; Ohta, Risa; Suga, Hiroaki

doi:10.1002/anie.201910894

Cited by 11 publications

(9 citation statements)

References 37 publications

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“…In principle, by modifying the γ-substitution of the 4bromovinylglycine derivative (propyl group in BrvG), this posttranslational chemical modification reaction is potentially applicable to the synthesis of peptides bearing more diverse azole structures, though it could be limited to those with substitutions at the 5th position. More importantly, this method can be integrated with other previously devised chemical posttranslational modification reactions [62][63][64][65][66][67] to yield peptides with various non-canonical backbone structures, expanding the chemical diversity of peptidomimetics accessible by in vitro translation.…”

Section: Resultsmentioning

confidence: 99%

Posttranslational chemical installation of azoles into translated peptides

et al. 2021

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Azoles are five-membered heterocycles often found in the backbones of peptidic natural products and synthetic peptidomimetics. Here, we report a method of ribosomal synthesis of azole-containing peptides involving specific ribosomal incorporation of a bromovinylglycine derivative into the nascent peptide chain and its chemoselective conversion to a unique azole structure. The chemoselective conversion was achieved by posttranslational dehydrobromination of the bromovinyl group and isomerization in aqueous media under fairly mild conditions. This method enables us to install exotic azole groups, oxazole and thiazole, at designated positions in the peptide chain with both linear and macrocyclic scaffolds and thereby expand the repertoire of building blocks in the mRNA-templated synthesis of designer peptides.

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Section: Resultsmentioning

confidence: 99%

Posttranslational chemical installation of azoles into translated peptides

et al. 2021

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“…82 In addition, the FIT−PatD system has been further integrated with a chemical modification to generate the Ψ[CH 2 NH] reduced amide backbone (Figure 9b). 83 The Ψ[CH 2 NH] reduced amide backbone is a well-documented peptide isostere and is widely used in bioactive peptidomimetics. Chemical reduction of the backbone thiazoline moiety produced in the FIT−PatD system resulted in thiazolidine, which could be further reduced to afford a Ψ[CH 2 NH] structure.…”

Section: Fit System Integrated With Ripp Enzymes For In Vitro Biosynt...mentioning

confidence: 99%

“…In addition, the FIT–PatD system has been further integrated with a chemical modification to generate the Ψ[CH 2 NH] reduced amide backbone (Figure b) . The Ψ[CH 2 NH] reduced amide backbone is a well-documented peptide isostere and is widely used in bioactive peptidomimetics.…”

Section: Combination Of the Fit System With Posttranslational Modific...mentioning

confidence: 99%

The RaPID Platform for the Discovery of Pseudo-Natural Macrocyclic Peptides

2021

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Conspectus Although macrocyclic peptides bearing exotic building blocks have proven their utility as pharmaceuticals, the sources of macrocyclic peptide drugs have been largely limited to mimetics of native peptides or natural product peptides. However, the recent emergence of technologies for discovering de novo bioactive peptides has led to their reconceptualization as a promising therapeutic modality. For the construction and screening of libraries of such macrocyclic peptides, our group has devised a platform to conduct affinity-based selection of massive libraries (>1012 unique sequences) of in vitro expressed macrocyclic peptides, which is referred to as the random nonstandard peptides integrated discovery (RaPID) system. The RaPID system integrates genetic code reprogramming using the FIT (flexible in vitro translation) system, which is largely facilitated by flexizymes (flexible tRNA-aminoacylating ribozymes), with mRNA display technology. We have demonstrated that the RaPID system enables rapid discovery of various de novo pseudo-natural peptide ligands for protein targets of interest. Many examples discussed in this Account prove that thioether-closed macrocyclic peptides (teMPs) obtained by the RaPID system generally exhibit remarkably high affinity and specificity, thereby potently inhibiting or activating a specific function(s) of the target. Moreover, such teMPs are used for a wide range of biochemical applications, for example, as crystallization chaperones for intractable transmembrane proteins and for in vivo recognition of specific cell types. Furthermore, recent studies demonstrate that some teMPs exhibit pharmacological activities in animal models and that even intracellular proteins can be inhibited by teMPs, illustrating the potential of this class of peptides as drug leads. Besides the ring-closing thioether linkage in the teMPs, genetic code reprogramming by the FIT system allows for incorporation of a variety of other exotic building blocks. For instance, diverse nonproteinogenic amino acids, hydroxy acids (ester linkage), amino carbothioic acid (thioamide linkage), and abiotic foldamer units have been successfully incorporated into ribosomally synthesized peptides. Despite such enormous successes in the conventional FIT system, multiple or consecutive incorporation of highly exotic amino acids, such as d- and β-amino acids, is yet challenging, and particularly the synthesis of peptides bearing non-carbonyl backbone structures remains a demanding task. To upgrade the RaPID system to the next generation, we have engaged in intensive manipulation of the FIT system to expand the structural diversity of peptides accessible by our in vitro biosynthesis strategy. Semilogical engineering of tRNA body sequences led to a new suppressor tRNA (tRNAPro1E2) capable of effectively recruiting translation factors, particularly EF-Tu and EF-P. The use of tRNAPro1E2 in the FIT system allows for not only single but also consecutive and multiple elongation of exotic amino acids, such as d-, β-, and γ-amino a...

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“…Previous work has shown that the cyclodehydratases PatD 14 and TruD 15 support leader sequence-dependent oxazoline/thiazoline formation within substrates containing nc-⍺-AAs adjacent to 16 or at the cyclization site itself. [17][18][19] In related work, it was shown that a chimeric leader peptide could direct the cyclodehydratase LynD 15 and the dehydrogenase TbtE 20 to install thiazol(in)es within substrates containing nc-⍺-AAs adjacent to the cyclization site. 21 Finally, reconstituted lactazole biosynthesis, 22 including the cyclodehydratase-dehydrogenase pair LazDE/LazF, was found to install oxazoles and thiazoles within polypeptide substrates containing ⍺-hydroxy, N-methyl, cyclic ⍺-, and β 3 -amino acids 23 at sites distal from the site of heterocyclization (> 4 residues away).…”

Section: Introductionmentioning

confidence: 99%

Redirecting RiPP biosynthetic enzymes to proteins and backbone-modified substrates

Walker¹,

Hamlish²,

Tytla³

et al. 2022

Preprint

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Ribosomally synthesized and post-translationally modified peptides (RiPPs) are peptide-derived natural products that include the FDA-approved analgesic ziconotide1,2 as well as compounds with potent antibiotic, antiviral, and anticancer properties.3 RiPP enzymes known as cyclodehydratases and dehydrogenases represent an exceptionally well-studied enzyme class.3 These enzymes work together to catalyze intramolecular, interresidue condensation3,4 and aromatization reactions that install oxazoline/oxazole and thiazoline/thiazole heterocycles within ribosomally produced polypeptide chains. Here we show that the previously reported enzymes MicD-F and ArtGox accept backbone-modified monomers, including aramids and beta-amino acids, within leader-free polypeptides, even at positions immediately preceding or following the site of cyclization/dehydrogenation. The products are sequence-defined chemical polymers with multiple, diverse, non-alpha-amino acid subunits. We show further that MicD-F and ArtGox can install heterocyclic backbones within protein loops and linkers without disrupting the native tertiary fold. Calculations reveal the extent to which these heterocycles restrict conformational space; they also eliminate a peptide bond. Both features could improve the stability or add function to linker sequences now commonplace in emerging biotherapeutics. Moreover, as thiazoles and thiazoline heterocycles are replete in natural products,5,6,7 small molecule drugs,8,9 and peptide-mimetic therapeutics,10 their installation in protein-based biotherapeutics could improve or augment performance, activity, stability, and/or selectivity. This work represents a general strategy to expand the chemical diversity of the proteome beyond and in synergy with what can now be accomplished by expanding the genetic code.

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Chemoenzymatic Posttranslational Modification Reactions for the Synthesis of Ψ[CH₂NH]‐Containing Peptides

Cited by 11 publications

References 37 publications

Posttranslational chemical installation of azoles into translated peptides

Posttranslational chemical installation of azoles into translated peptides

The RaPID Platform for the Discovery of Pseudo-Natural Macrocyclic Peptides

Redirecting RiPP biosynthetic enzymes to proteins and backbone-modified substrates

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Chemoenzymatic Posttranslational Modification Reactions for the Synthesis of Ψ[CH2NH]‐Containing Peptides

Cited by 11 publications

References 37 publications

Posttranslational chemical installation of azoles into translated peptides

Posttranslational chemical installation of azoles into translated peptides

The RaPID Platform for the Discovery of Pseudo-Natural Macrocyclic Peptides

Redirecting RiPP biosynthetic enzymes to proteins and backbone-modified substrates

Contact Info

Product

Resources

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Chemoenzymatic Posttranslational Modification Reactions for the Synthesis of Ψ[CH₂NH]‐Containing Peptides