Department of Biochemistry and Molecular Pharmacology; RNA Therapeutics Institute
Amino Acid Sequence; Amino Acid Substitution; Color; Fluorescence Resonance Energy Transfer; Genes, Reporter; HeLa Cells; Humans; Luminescent Proteins; Molecular Probes; Molecular Sequence Data; Mutation; Photochemistry; Sequence Alignment; Sequence Homology
Biochemistry | Biochemistry, Biophysics, and Structural Biology | Chemistry | Molecular Biology
We used a red chromophore formation pathway, in which the anionic red chromophore is formed from the neutral blue intermediate, to suggest a rational design strategy to develop blue fluorescent proteins with a tyrosine-based chromophore. The strategy was applied to red fluorescent proteins of the different genetic backgrounds, such as TagRFP, mCherry, HcRed1, M355NA, and mKeima, which all were converted into blue probes. Further improvement of the blue variant of TagRFP by random mutagenesis resulted in an enhanced monomeric protein, mTagBFP, characterized by the substantially higher brightness, the faster chromophore maturation, and the higher pH stability than blue fluorescent proteins with a histidine in the chromophore. The detailed biochemical and photochemical analysis indicates that mTagBFP is the true monomeric protein tag for multicolor and lifetime imaging, as well as the outstanding donor for green fluorescent proteins in Forster resonance energy transfer applications.
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Copyright 2008 Elsevier Ltd. Publisher pdf posted as allowed by publisher's Elsevier user licence at http://www.elsevier.com/about/open-access/open-access-policies/oa-license-policy/elsevier-user-license.
DOI of Published Version
Subach OM, Gundorov IS, Yoshimura M, Subach FV, Zhang J, Grüenwald D, Souslova EA, Chudakov DM, Verkhusha VV. Conversion of red fluorescent protein into a bright blue probe. Chem Biol. 2008 Oct 20;15(10):1116-24. doi:10.1016/j.chembiol.2008.08.006. Link to article on publisher's site
Chemistry and biology
Subach, Oksana M.; Gundorov, Illia S.; Yoshimura, Masami; Subach, Fedor V.; Zhang, Jinghang; Grunwald, David; Souslova, Ekaterina A.; Chudakov, Dmitriy M.; and Verkhusha, Vladislav V., "Conversion of red fluorescent protein into a bright blue probe" (2008). Biochemistry and Molecular Pharmacology Publications and Presentations. 171.