Screening of 2A peptides for polycistronic gene expression in yeast
Journal article, 2018

A complexity of pathway expression in yeast compared to prokaryotes is the need for separate promoters and terminators for each gene expressed. Single transcript expression and separated protein production is possible via the use of 2A viral peptides, but detailed characterization to assess their suitability and applications is needed. The present work aimed to characterize multiple 2A peptide sequences to determine suitability for metabolic engineering applications in Saccharomyces cerevisiae. We screened 22 peptides placed between fluorescent protein sequences. Cleaving efficiency was calculated by western blot intensity of bands corresponding to the cleaved and uncleaved forms of the reporter. Three out of the 22 sequences showed high cleavage efficiency: 2A peptide from Equine rhinitis B virus (91%), Porcine teschovirus-1 (85%) and Operophtera brumata cypovirus-18 (83%). Furthermore, expression of the released protein was comparable to its monocistronic expression. As a proof-of-concept, the triterpene friedelin was successfully produced in the same yeast strain by expressing its synthase with the truncated form of HMG1 linked by the 2A peptide of ERBV-1, with production titers comparable to monocistronic expression (via separate promoters). These results suggest that these peptides could be suitable for expression and translation of multiple proteins in metabolic engineering applications in S. cerevisiae.

stop-carry on

self-cleavage

ERBV-1 2A peptide

Saccharomyces cerevisiae

Yeast metabolic engineering

polycistronic

Multi-gene expression

Author

Tatiana M. Souza-Moreira

São Paulo State University (UNESP)

Clara Navarrete Roman

Chalmers, Biology and Biological Engineering, Systems and Synthetic Biology

Xin Chen

Chalmers, Biology and Biological Engineering, Systems and Synthetic Biology

Cleslei F. Zanelli

São Paulo State University (UNESP)

Sandro R. Valentini

São Paulo State University (UNESP)

Maysa Furlan

São Paulo State University (UNESP)

Jens B Nielsen

Technical University of Denmark (DTU)

Chalmers, Biology and Biological Engineering, Systems and Synthetic Biology

Anastasia Krivoruchko

Biopetrolia

Chalmers, Biology and Biological Engineering, Systems and Synthetic Biology

FEMS Yeast Research

1567-1356 (ISSN) 1567-1364 (eISSN)

Vol. 18 5 foy036

Subject Categories

Biochemistry and Molecular Biology

Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)

Other Industrial Biotechnology

DOI

10.1093/femsyr/foy036

More information

Latest update

12/10/2018