An Evolutionary Strategy for Identification of Higher Order, Green Fluorescent Host–Guest Pairs Compatible with Living Systems

Garrett R. Casey, Xinqi Zhou, Lauren Lesiak, Bi Xu, Yuan Fang, Donald F. Becker, Cliff I. Stains

Research output: Contribution to journalArticlepeer-review

1 Scopus citations

Abstract

Engineered miniprotein host–small-molecule guest pairs could be utilized to design new processes within cells as well as investigate fundamental aspects of cell signaling mechanisms. However, the development of host–guest pairs capable of functioning in living systems has proven challenging. Moreover, few examples of host–guest pairs with stoichiometries other than 2:1 exist, significantly hindering the ability to study the influence of oligomerization state on signaling fidelity. Herein, we present an approach to identify host–guest systems for relatively small green fluorescent guests by incorporation into cyclic peptides. The optimal host–guest pair produced a 10-fold increase in green fluorescence signal upon binding. Biophysical characterization clearly demonstrated higher order supramolecular assembly, which could be visualized on the surface of living yeast cells using a turn-on fluorescence readout. This work further defines evolutionary design principles to afford host–guest pairs with stoichiometries other than 2:1 and enables the identification of spectrally orthogonal host–guest pairs.

Original languageEnglish (US)
Pages (from-to)16721-16726
Number of pages6
JournalChemistry - A European Journal
Volume26
Issue number70
DOIs
StatePublished - Dec 15 2020

Keywords

  • directed evolution
  • fluorescent probes
  • host–guest systems
  • protein engineering
  • self-assembly

ASJC Scopus subject areas

  • General Chemistry
  • Catalysis
  • Organic Chemistry

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