Recombinant Expression and Phenotypic Screening of a Bioactive Cyclotide Against α-Synuclein-Induced Cytotoxicity in Baker′s Yeast

Krishnappa Jagadish, Andrew Gould, Radhika Borra, Subhabrata Majumder, Zahid Mushtaq, Alexander Shekhtman, Julio A. Camarero

Research output: Contribution to journalArticlepeer-review

37 Scopus citations

Abstract

We report for the first time the recombinant expression of fully folded bioactive cyclotides inside live yeast cells by using intracellular protein trans-splicing in combination with a highly efficient split-intein. This approach was successfully used to produce the naturally occurring cyclotide MCoTI-I and the engineered bioactive cyclotide MCoCP4. Cyclotide MCoCP4 was shown to reduce the toxicity of human α-synuclein in live yeast cells. Cyclotide MCoCP4 was selected by phenotypic screening from cells transformed with a mixture of plasmids encoding MCoCP4 and inactive cyclotide MCoTI-I in a ratio of 1:5×104. This demonstrates the potential for using yeast to perform phenotypic screening of genetically encoded cyclotide-based libraries in eukaryotic cells.

Original languageEnglish (US)
Pages (from-to)8390-8394
Number of pages5
JournalAngewandte Chemie - International Edition
Volume54
Issue number29
DOIs
StatePublished - Jul 13 2015

Keywords

  • aptamers
  • biological activity
  • peptides
  • protein expression
  • synuclein

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