A sensitive and specific genetically-encoded potassium ion biosensor for in vivo applications across the tree of life

Abstract

Potassium ion (K+) plays a critical role as an essential electrolyte in all biological systems. Genetically encoded fluorescent K+ biosensors are promising tools to further improve our understanding of K+-dependent processes under normal and pathological conditions. Here, we report the crystal structure of a previously reported genetically encoded fluorescent K+ biosensor, GINKO1, in the K+-bound state. Using structure-guided optimization and directed evolution, we have engineered an improved K+ biosensor, designated GINKO2, with higher sensitivity and specificity. We have demonstrated the utility of GINKO2 for in vivo detection and imaging of K+ dynamics in multiple model organisms, including bacteria, plants, and mice.

Adaptable, Turn-On Monobody (ATOM) Fluorescent Biosensors for Multiplexed Detection in Cells

“…If our results are representative, at least one of the proteins will likely be a functioning ATOM sensor. This process stands in contrast to the large libraries and multiple iterations of screening and/or selection that are employed when developing many other single-molecule fluorescent biosensors 34,34,35 .…”

Modern analytical and bioanalytical technologies and concepts for smart and precision farming

“…86 The performance of biosensors was further evaluated on live organisms such as bacteria, plants, and rats. 87 The result revealed the ability of the bioristor to function as a potassium indicator without inhibiting plant growth.…”

Fluorescent proteins and genetically encoded biosensors

“…1 in ref. 257). Similarly, GEPII makes an insertion of Kbp into the linkage between mseCFP and cpV (CFP and Venus/YFP mutants), and its FRET is activated when Kbp closes its lobes in K + -bound form.…”

“…Both series use a same potassiumbinding protein (Kbp) from E. coli, which has a BON domain and a LysM domain, hinged by flexible loops, which makes the lobeclose action when bound to a K + ion. The sensors, namely GEPII 255 GINKO, 256,257 are FRET-based and single-FP based, respectively. In GINKO, the EGFP secondary structures are not permuted, but make a simple insertion of two Kbp lobes at the Tyr145 site.…”