Recombinant Expression and Phenotypic Screening of a Bioactive Cyclotide Against α-Synuclein-Induced Cytotoxicity in Baker's Yeast.
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 1:5×10(4). This demonstrates the potential for using yeast to perform phenotypic screening of genetically encoded cyclotide-based libraries in eukaryotic cells.
['Jagadish|Krishnappa|K|', 'Gould|Andrew|A|', 'Borra|Radhika|R|', 'Majumder|Subhabrata|S|', 'Mushtaq|Zahid|Z|', 'Shekhtman|Alexander|A|', 'Camarero|Julio A|JA|']
D000595:Amino Acid Sequence D047169:Cyclotides D006801:Humans D008958:Models, Molecular D008969:Molecular Sequence Data D015202:Protein Engineering D011994:Recombinant Proteins D012441:Saccharomyces cerevisiae D020040:Trans-Splicing D014170:Transformation, Genetic D051844:alpha-Synuclein
aptamers biological activity peptides protein expression synuclein