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Enhanced Photostability of Genetically Encodable Fluoromodules Based on Fluorogenic Cyanine Dyes and a Promiscuous Protein Partner

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journal contribution
posted on 16.09.2009, 00:00 by Nathaniel I. Shank, Kimberly J. Zanotti, Frederick Lanni, Peter B. Berget, Bruce A. Armitage
Fluoromodules are discrete complexes of biomolecules and fluorogenic dyes. Binding of the dyes to their cognate biomolecule partners results in enhanced dye fluorescence. We exploited a previously reported promiscuous binding interaction between a single-chain, variable fragment antibody protein and a family of cyanine dyes to create new protein−dye fluoromodules that exhibit enhanced photostability while retaining high affinity protein−dye binding. Modifications to the dye structure included electron-withdrawing groups that provide resistance to photo-oxidative damage. Low nanomolar equilibrium dissociation constants were found for the new dyes. Fluorescence microscopy illustrates how yeast can be surface-labeled with three different colors based on a single protein and appropriately chosen dyes.