Live‐Cell Copper‐Induced Fluorescence Quenching of the Flavin‐Binding Fluorescent Protein CreiLOV. (21st January 2020)

• Record Type: Journal Article
• Authors: Zou, Wenping; Le, Khoa; Zastrow, Melissa L.
• Is Part Of: Chembiochem. Volume 21:Number 9(2020)
• Page Start: 1356
• Page End: 1363
• Publication Date: 2020-01-21
• Subjects: bacteria -- copper -- fluorescence lifetime -- fluorescent proteins -- metal sensors; Biochemistry -- Periodicals; Molecular biology -- Periodicals; Pharmaceutical chemistry -- Periodicals; 572
• DOI: 10.1002/cbic.201900669
• Languages: English
• ISSNs: 1439-4227

Abstract

Abstract: CreiLOV is a flavin‐binding fluorescent protein derived from the blue‐light photoreceptor protein family that contains light‐oxygen‐voltage (LOV) sensing domains. Flavin‐binding fluorescent proteins represent a promising foundation for new fluorescent reporters and biosensors that can address limitations of the well‐established green fluorescent protein (GFP) family. Flavin‐binding fluorescent proteins are smaller than GFPs, are stable over a wider pH range, offer rapid chromophore incorporation, and are oxygen‐independent so can be applied to live anaerobic organisms. Among the flavin‐binding fluorescent proteins, CreiLOV has a high quantum yield and excellent photophysical properties, making it promising for cellular applications. Here, we investigated the suitability of CreiLOV as an intensity‐ and fluorescence‐lifetime‐based metal sensor. CreiLOV selectively binds copper(II) over other biologically relevant metals with low‐micromolar affinity, resulting in fluorescence quenching and a decrease in the fluorescence lifetime that can be observed in cuvettes and live bacterial cells. Abstract : In‐depth reporting : Flavin‐binding proteins are promising fluorescent reporters in metal sensing that do not rely on oxygen, offer rapid chromophore incorporation, and are small. Here we show that CreiLOV selectively detects copper (II) over other biological metals in vitro and in live cells by using fluorescence intensity and lifetime‐based detection methods.