Triazene Coarctate Cyclization for the Synthesis of Fluorescent Cyclic Peptides Restricted; Files Only

Abstract

Fluorescent cyclic peptides have transformed the way we study biological systems by tracking interactions with proteins and other biomolecules in situ or imaging biological events in real-time with high spatial resolution. Current methods mainly rely on functionalizing side chains of cyclic peptides with fluorophores, which could change the physicochemical properties of the parent cyclic peptides and influence their ability to bind and localize in cells. None of the current methods are capable of generating cyclic peptides with in-built fluorescence for cell imaging studies. Herein, we introduce a simple, chemoselective triazene coarctate cyclization (TCC) reaction that combines N-terminal proline or methylated lysine with ortho-alkyne functionalized phenyl diazonium ions to generate highly stable, fluorescent isoindazole 3-carbaldehyde peptides. The peptides exhibit fluorescence only after formation of the isoindazole 3-carbaldehyde, and the photophysical properties of the product can be tuned through late-stage modifications of the in-built aldehyde. We envision the application of this method to cellular imaging in both UV and IR regions. The scope of this reaction is broad, opening up a powerful new approach for the synthesis of in-built fluorescent cyclic peptides and for diverse applications in biological systems. 

Table of Contents

Introduction. 1

Results and discussion. 5

Chapter 1: Background, inspiration, and approach. 5

Chapter 2: Synthesis of p-amino-m-ethynyl phenylalanine-containing peptides. 8

Chapter 3: Converting linear peptides to cyclic triazenes. 14

Chapter 4: Coarctate chemistry to convert cyclic triazenes to isoindazole 3-carbaldehyde peptides. 17

Chapter 5: Derivatization of the isoindazole 3-carbaldehyde moiety. 23

Conclusions and future directions. 29

Acknowledgement of contributions. 31

References. 32

Supplemental Information. 35

General 35

Materials. 35

Purification. 35

Instrumentation and sample analysis. 36

Fmoc Solid-Phase Peptide Synthesis (Fmoc-SPPS) 36

Supplementary Figure 1: Synthesis of Fmoc-Phe(4-NH2,3-I)-OH.. 37

Supplementary Figure 2: Synthesis of Fmoc-Phe(4-NH2,3-(trimethylsilyl)ethyne)-OH.. 40

Supplementary Figure 3: Solid-phase Sonogashira coupling. 43

Supplementary Figure 4: Triazene peptide synthesis. 45

Supplementary Figure 5: Optimization of the coarctate reaction on peptides. 48

Supplementary Figure 6: Synthesis of small molecule triazene 11. 52

Supplementary Figure 7: Optimization of small molecule isoindazole 3-carbaldehyde 12 Synthesis 53

Supplementary Figure 8: Synthesis of Wittig, Horner-Wittig, and Henry Olefination Products. 57

Supplementary Figure 9: Mechanistic considerations for formation of aryl diazonium and triazene. 79

Supplementary Figure 10: Mechanisms for derivatization of aldehyde from Figure 14. 81

Supplementary Figure 11: Amino acid coupling mechanism and discussion. 82

References 83

About this Honors Thesis

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School	Emory College
Department	Chemistry
Degree	B.S.
Submission	Honors Thesis
Language	English
Research Field	Chemistry, Biochemistry Chemistry, Organic Chemistry, Polymer
Parola chiave	Fluorescence Organic Synthesis Peptide Cyclization Fluorescent Peptides Cyclic Peptides Peptides Amino Acids
Committee Chair / Thesis Advisor	Monika Raj, Emory University
Committee Members	Matthew Weinschenk, Emory University Tracy McGill, Emory University Richard Himes, Emory University Elena Cholakova, Emory University

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