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TitleEngineering of mCherry variants with long Stokes shift, red-shifted fluorescence, and low cytotoxicity.
Publication TypeJournal Article
Year of Publication2017
AuthorsShen, Yi, Yingche Chen, Jiahui Wu, Nathan C. Shaner, and Robert E. Campbell
JournalPLoS One
Volume12
Issue2
Paginatione0171257
Date Published2017
ISSN1932-6203
KeywordsAnimals, Anthozoa, Escherichia coli, Evolution, Molecular, Fluorescence, Fluorescent Dyes, Gene Library, Luminescent Proteins, Models, Molecular, Mutagenesis, Site-Directed, Oligonucleotides, Protein Engineering
Abstract

MCherry, the Discosoma sp. mushroom coral-derived monomeric red fluorescent protein (RFP), is a commonly used genetically encoded fluorophore for live cell fluorescence imaging. We have used a combination of protein design and directed evolution to develop mCherry variants with low cytotoxicity to Escherichia coli and altered excitation and emission profiles. These efforts ultimately led to a long Stokes shift (LSS)-mCherry variant (λex = 460 nm and λem = 610 nm) and a red-shifted (RDS)-mCherry variant (λex = 600 nm and λem = 630 nm). These new RFPs provide insight into the influence of the chromophore environment on mCherry's fluorescence properties, and may serve as templates for the future development of fluorescent probes for live cell imaging.

DOI10.1371/journal.pone.0171257
Alternate JournalPLoS ONE
PubMed ID28241009
PubMed Central IDPMC5328254