PT  - JOURNAL ARTICLE
AU  - Parashar Thapa
AU  - Robert Stewart
AU  - Rebecka J. Sepela
AU  - Oscar Vivas
AU  - Laxmi K. Parajuli
AU  - Mark Lillya
AU  - Sebastian Fletcher-Taylor
AU  - Bruce E. Cohen
AU  - Karen Zito
AU  - Jon T. Sack
TI  - Optical measurement of endogenous ion channel voltage sensor activation in tissue
AID  - 10.1101/541805
DP  - 2019 Jan 01
TA  - bioRxiv
PG  - 541805
4099  - http://biorxiv.org/content/early/2019/09/05/541805.short
4100  - http://biorxiv.org/content/early/2019/09/05/541805.full
AB  - A primary goal of molecular physiology is to understand how conformational changes of proteins affect the function of cells, tissues, and organisms. Here, we describe an imaging method for measuring the conformational changes of a voltage-sensing protein within tissue. We synthesized a fluorescent molecular probe, compatible with two-photon microscopy, that targets a resting conformation of Kv2-type voltage gated K+ channel proteins. Voltage-response characteristics were used to calibrate a statistical thermodynamic model relating probe labeling intensity to the conformations adopted by unlabeled Kv2 proteins. Two-photon imaging of rat brain slices labeled with the probe revealed fluorescence consistent with conformation-selective labeling of endogenous neuronal Kv2 proteins. In principle, this method of quantifying endogenous protein conformational change from fluorescence images is generalizable to other proteins labeled with conformation-selective probes.