Application of Zinpyr-1 for the investigation of zinc signals in Escherichia coli

Changes of the pico- to nanomolar concentration of free intracellular Zn2+ are part of the signal transduction in mammalian cells. These zinc signals regulate the enzymatic activity of target proteins such as protein tyrosine phosphatases. For Escherichia coli, previous studies have reported diverging concentrations from femto- to picomolar, raising the question if Zn2+ could also have a function in bacterial signaling. This manuscript explores the use of the low molecular weight fluorescent probe Zinpyr-1 in E. coli. The probe detects free Zn2+ in these bacteria. Comparable to mammalian cells, other metal ions, especially Hg2+ and Cd2+, interfere with the detection of Zn2+. Moreover, experiments in E. coli were particularly prone to artifacts based on cellular autofluorescence, necessitating corrections that are not required in mammalian cells. Based on measurements in lysates of E. coli and the mammalian cell line Jurkat, similar values between 0.1 and 0.2 nM free Zn2+ were found. For E. coli, this corresponds to less than one free zinc ion per cell. Moreover, phosphatase inhibition by Zn2+ was only observed in Jurkat, but not E. coli. This excludes a function for zinc signals as a regulator of bacterial phosphatases. Still, changes in the free Zn2+ concentration were observed in response to elevated extracellular Zn2+ and pH, or to addition of the detergent NP-40, suggesting that other processes could be controlled by the free intracellular Zn2+ concentration.