Rapid Fluorescence Assay for Polyphosphate in Yeast Extracts Using JC-D7.

Polyphosphate (polyP) is an intriguing molecule that is found in almost any organism, covering a multitude of cellular functions. In industry, polyP is used due to its unique physiochemical properties, including pH buffering, water binding, and bacteriostatic activities. Despite the importance of polyP, its analytics is still challenging, with the gold standard being 31P NMR. Here, we present a simple staining method using the fluorescent dye JC-D7 for the semi-quantitative polyP evaluation in yeast extracts. Notably, fluorescence response was affected by polyP concentration and polymer chain length in the 0.5-500 µg/mL polyP concentration range. Hence, for polyP samples of unknown chain compositions, JC-D7 cannot be used for absolute quantification. Fluorescence of JC-D7 was unaffected by inorganic phosphate up to 50 mM. Trace elements (FeSO4 > CuSO4 > CoCl2 > ZnSO4) and toxic mineral salts (PbNO3 and HgCl2) diminished polyP-induced JC-D7 fluorescence, affecting its applicability to samples containing polyP-metal complexes. The fluorescence was only marginally affected by other parameters, such as pH and temperature. After validation, this simple assay was used to elucidate the degree of polyP production by yeast strains carrying gene deletions in (poly)phosphate homeostasis. The results suggest that staining with JC-D7 provides a robust and sensitive method for detecting polyP in yeast extracts and likely in extracts of other microbes. The simplicity of the assay enables high-throughput screening of microbes to fully elucidate and potentially enhance biotechnological polyP production, ultimately contributing to a sustainable phosphorus utilization.

Deitert A ，Fees J ，Mertens A ，Nguyen Van D ，Maares M ，Haase H ，Blank LM ，Keil C ... -  《-》

Quantification of Polyphosphate in Environmental Planktonic Samples Using a Novel Fluorescence Dye JC-D7.

Yang X ，Gao R ，Zhang Q ，Yung CCM ，Yin H ，Li J ... -  《-》

Dependence of inorganic polyphosphate chain length on the orthophosphate content in the culture medium of the yeast Saccharomyces cerevisiae.

Vagabov VM ，Trilisenko LV ，Kulaev IS 《BIOCHEMISTRY-MOSCOW》

Vtc5 Is Localized to the Vacuole Membrane by the Conserved AP-3 Complex to Regulate Polyphosphate Synthesis in Budding Yeast.

Bentley-DeSousa A ，Downey M 《mBio》

Direct labeling of polyphosphate at the ultrastructural level in Saccharomyces cerevisiae by using the affinity of the polyphosphate binding domain of Escherichia coli exopolyphosphatase.

Saito K ，Ohtomo R ，Kuga-Uetake Y ，Aono T ，Saito M ... -  《-》