Dempski, Robert E.
Channelrhodopsin-2 (ChR2) is a photoactive cation channel permeable to mono and divalent ions. The max response of wild-type ChR2 occurs at 470 nm light (λmax). ChR2 can be used to map neuronal networks. We hypothesized mutations at G181, would alter the electrostatic environment of the retinal binding pocket and alter λmax. G was mutated to S, T, K, D and E. Action spectra were recorded by measuring max current responses with two-electrode voltage clamp as a function of wavelength with tunable bandpass filters. G181S and G181T had little to no effect on λmax but had reduced function. All charged mutants had no activation, a consequence of surface expression shown by Western blotting. Therefore, we suggest that G181 is important for the proper surface expression of ChR2.
Worcester Polytechnic Institute
Major Qualifying Project
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Chemistry and Biochemistry