Synthesis of Novel Peptide Linkers: Simultaneous Cyclization and Labeling

Dewkar, Gajanan K.; Carneiro, Pedro B.; Hartman, Matthew C. T.

doi:10.1021/ol901662c

Cited by 16 publications

(17 citation statements)

References 24 publications

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“…We and others (Dewkar, Carneiro, & Hartman, 2009; Timmerman et al, 2005) have shown that the peptide concentration for this reaction is optimal between 0.01 and 1 m M , but can also be achieved at higher concentrations. One of the byproducts encountered if the peptide is too concentrated is the cross-linked dimer.…”

Section: Applications Tips and Troubleshootingmentioning

confidence: 75%

Conformational Restriction of Peptides Using Dithiol Bis-Alkylation

Peraro

Siegert

Kritzer

2016

Methods in Enzymology

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Macrocyclic peptides are highly promising as inhibitors of protein–protein interactions. While many bond-forming reactions can be used to make cyclic peptides, most have limitations that make this chemical space challenging to access. Recently, a variety of cysteine alkylation reactions have been used in rational design and library approaches for cyclic peptide discovery and development. We and others have found that this chemistry is versatile and robust enough to produce a large variety of conformationally constrained cyclic peptides. In this chapter, we describe applications, methods, mechanistic insights, and troubleshooting for dithiol bis-alkylation reactions for the production of cyclic peptides. This method for efficient solution-phase macrocyclization is highly useful for the rapid production and screening of loop-based inhibitors of protein–protein interactions.

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Section: Applications Tips and Troubleshootingmentioning

confidence: 75%

Conformational Restriction of Peptides Using Dithiol Bis-Alkylation

Peraro

Siegert

Kritzer

2016

Methods in Enzymology

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“…For cyclization, we focused on the well-established and robust bis-bromomethylbenzene chemistry, which covalently cyclizes peptides with two cysteines. 33–35 The random position of the second cysteine allows the BRCA1 protein to “choose” from a variety of ring sizes or linear peptides for an optimum fit. Based on our previous work describing amino acid analogues that can be incorporated by substitution in ribosomal translation, 36 a group of six unnatural amino acids (Figure 1, Supporting Information Chart S1) were optimized for use together with the other 14 natural amino acids to make up the building blocks for the peptide selection (Supporting Information Figures S1–S5 and Table S1).…”

Section: Resultsmentioning

confidence: 99%

Peptide Library Approach to Uncover Phosphomimetic Inhibitors of the BRCA1 C-Terminal Domain

White

Sun

Ma³

et al. 2015

ACS Chem. Biol.

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Many intracellular protein–protein interactions are mediated by the phosphorylation of serine, and phosphoserine-containing peptides can inhibit these interactions. However, hydrolysis of the phosphate by phosphatases, and the poor cell permeability associated with phosphorylated peptides has limited their utility in cellular and in vivo contexts. Compounding the problem, strategies to replace phosphoserine in peptide inhibitors with easily accessible mimetics (such as Glu or Asp) routinely fail. Here, we present an in vitro selection strategy for replacement of phosphoserine. Using mRNA display, we created a 10 trillion member structurally diverse unnatural peptide library. From this library, we found a peptide that specifically binds to the C-terminal domain (BRCT)2 of breast cancer associated protein 1 (BRCA1) with an affinity comparable to phosphorylated peptides. A crystal structure of the peptide bound reveals that the pSer-x-x-Phe motif normally found in BRCA1 (BRCT)2 binding partners is replaced by a Glu-x-x-4-fluoroPhe and that the peptide picks up additional contacts on the protein surface not observed in cognate phosphopeptide binding. Expression of the peptide in human cells led to defects in DNA repair by homologous recombination, a process BRCA1 is known to coordinate. Overall, this work validates a new in vitro selection approach for the development of inhibitors of protein–protein interactions mediated by serine phosphorylation.

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“…23 Inhibitors of the p53-HDM2 interaction reactivate p53 and are expected to induce apoptosis to malignant cells. 24 X-ray crystallographic study of the complex of peptide p53 [15][16][17][18][19][20][21][22][23][24][25][26][27][28][29] (SQETFSDLWKLLPEN), derived from the transactivation domain of p53, with HDM2 revealed that p53 [15][16][17][18][19][20][21][22][23][24][25][26][27][28][29] formed a ahelical structure in binding and the hydrophobic residues (Phe19/Trp23/Leu26) interacted with the cleft on HDM2. 25 Peptide inhibitors and peptide mimetic compounds have been developed for regulating the p53-HDM2 interaction.…”

Section: Design Of Inhibitory Peptides Based On Yt1-s Analog For P53-mentioning

confidence: 99%

“…Cyclization methods with α,α′‐dibromoxylene and the derivatives have been successfully developed to constrain peptides . Although simultaneous cyclization and labeling of peptides with 1,3,5‐tris(bromomethyl)benzene was achieved, it required monoalkylation of the precursor linker before simultaneous cyclization and labeling . To develop a feasible method for introducing a variety of functional molecules to the cyclized peptides by using the CuAAC reaction, here we used 1,3‐di(bromomethyl)‐5‐propargyloxybenzene ( DBMPB ), originally synthesized for the intermediate to produce a linker for hyperbranched polymers .…”

Section: Introductionmentioning

confidence: 99%

“…[17][18][19] Although simultaneous cyclization and labeling of peptides with 1,3,5-tris(bromomethyl)benzene was achieved, it required monoalkylation of the precursor linker before simultaneous cyclization and labeling. 20 To develop a feasible method for introducing a variety of functional molecules to the cyclized peptides by using the CuAAC reaction, 21 here we used 1,3di(bromomethyl)-5-propargyloxybenzene (DBMPB), originally synthesized for the intermediate to produce a linker for hyperbranched polymers. 22 Functional molecules with azide can react with the linkage site via a propargyl group in the CuAAC reaction.…”

Section: Introductionmentioning

confidence: 99%

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Macrocyclization and labeling of helix–loop–helix peptide with intramolecular bis‐thioether linkage

et al. 2016

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Conformationally constrained peptides have been developed as an inhibitor for protein-protein interactions (PPIs), and we have de novo designed cyclized helix-loop-helix (cHLH) peptide with a disulfide bond consisting of 40 amino acids to generate molecular-targeting peptides. However, synthesis of long peptides has sometimes resulted in low yield according to the respective amino acid sequences. Here we developed a method for efficient synthesis and labeling for cHLH peptides. First, we synthesized two peptide fragments and connected them by the copper-mediated alkyne and azide cycloaddition (CuAAC) reaction. Cyclization was performed by bis-thioether linkage using 1,3-dibromomethyl-5-propargyloxybenzene, and subsequently, the cHLH peptide was labeled with an azide-labeled probe. Finally, we designed and synthesized a peptide inhibitor for the p53-HDM2 interaction using a structure-guided design and successfully labeled it with a fluorescent probe or a functional peptide, respectively, by click chemistry. This macrocyclization and labeling method for cHLH peptide would facilitate the discovery of de novo bioactive ligands and therapeutic leads. © 2016 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 106: 415-421, 2016.

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Synthesis of Novel Peptide Linkers: Simultaneous Cyclization and Labeling

Cited by 16 publications

References 24 publications

Conformational Restriction of Peptides Using Dithiol Bis-Alkylation

Conformational Restriction of Peptides Using Dithiol Bis-Alkylation

Peptide Library Approach to Uncover Phosphomimetic Inhibitors of the BRCA1 C-Terminal Domain

Macrocyclization and labeling of helix–loop–helix peptide with intramolecular bis‐thioether linkage

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