Sung, Uhna Sepehri-Rad, Masoud Piao, Hong Hua Jin, Lei Hughes, Thomas Cohen, Lawrence B. Baker, Bradley J. 2024-01-20T05:33:11Z 2024-01-20T05:33:11Z 2021-09-03 2015-11-20 1932-6203 https://pubs.kist.re.kr/handle/201004/124739 FRET (Forster Resonance Energy Transfer)-based protein voltage sensors can be useful for monitoring neuronal activity in vivo because the ratio of signals between the donor and acceptor pair reduces common sources of noise such as heart beat artifacts. We improved the performance of FRET based genetically encoded Fluorescent Protein (FP) voltage sensors by optimizing the location of donor and acceptor FPs flanking the voltage sensitive domain of the Ciona intestinalis voltage sensitive phosphatase. First, we created 39 different "Nabi1" constructs by positioning the donor FP, UKG, at 8 different locations downstream of the voltage-sensing domain and the acceptor FP, mKO, at 6 positions upstream. Several of these combinations resulted in large voltage dependent signals and relatively fast kinetics. Nabi1 probes responded with signal size up to 11%Delta F/F for a 100 mV depolarization and fast response time constants both for signal activation (similar to 2ms) and signal decay (similar to 3ms). We improved expression in neuronal cells by replacing the mKO and UKG FRET pair with Clover (donor FP) and mRuby2 (acceptor FP) to create Nabi2 probes. Nabi2 probes also had large signals and relatively fast time constants in HEK293 cells. In primary neuronal culture, a Nabi2 probe was able to differentiate individual action potentials at 45 Hz. English PUBLIC LIBRARY SCIENCE ACTION-POTENTIALS OPTICAL ELECTROPHYSIOLOGY SENSING PHOSPHATASE ELECTRICAL-ACTIVITY DYNAMIC-RANGE NEURONS DOMAIN GREEN DIMERIZATION SPIKING Developing Fast Fluorescent Protein Voltage Sensors by Optimizing FRET Interactions Article 10.1371/journal.pone.0141585 1 PLOS ONE, v.10, no.11 PLOS ONE 10 11 scie scopus 000365251500004 2-s2.0-84957990849 Multidisciplinary Sciences Science & Technology - Other Topics Article ACTION-POTENTIALS OPTICAL ELECTROPHYSIOLOGY SENSING PHOSPHATASE ELECTRICAL-ACTIVITY DYNAMIC-RANGE NEURONS DOMAIN GREEN DIMERIZATION SPIKING