Solubilization and partial characterization of the tetrodotoxin binding component from electric eel electroplaque

Citation

Stackhouse, Frank A. (1977) Solubilization and partial characterization of the tetrodotoxin binding component from electric eel electroplaque. Master's thesis, California Institute of Technology. doi:10.7907/EGBS-9230. https://resolver.caltech.edu/CaltechTHESIS:04282010-093315226

Abstract

The tetrodotoxin (TTX) binding component from the electroplaque of Electrophorus electricus was solubilized and studied using ^3H-TTX purified by a modified procedure using Bio-Rex-70 at pH 8.7. The best extraction yield was obtained from a solubilization in 1-2% Lubroi-PX, at pH 7.6, for 2 hours, at a membrane fragment concentration equivalent to 2.0 - 2.5 g original wet weight tissue/ml. Gel filtration on Sepharose-6B was used to verify that the TTX binding component was solubilized. The molecular weight of the solubilized component was found to be about 200,000 using 10-43% glycerol gradients. A Sephadex G-50 assay was used to study binding. A K_D of 1.2 nM was obtained for ^3H-TTX binding. This binding was affected by cations in a manner similar to that found by Reed and Raftery [Biochemistry 15:944-953 (1975)]. The solubilized material was found to be unstable with respect to TTX binding; this stability was affected by the presence of cations and TTX, but not veratridine or procaine.

Thesis Files