Purification from rat sarcolemma of component of the excitable membrar (neurotoxin/ion channel/muscle/action potential/glycoprotein affi)

The saxitoxin-binding component (SBC) of the excitable membrane sodium channel has been solubilized and 1 purified from rat skeletal muscle sarcolemma. Phospholipid was 1 required in mixed micelles with detergent for stability of the mammalian SBC. Even at optimal detergent-to-phospholipid ) ratio, the solubilized SBC showed significant temperature- dependent loss of specific toxin binding with time, necessitating maintenance of low temperatures during purification. Char- < acteristics of saxitoxin binding to the solubilized material closely resembled those seen in intact membranes. A weak anion-ex- change column was synthesized; it provided rapid 10- to 20-fold purification of the solubilized SBC. Additional necessary pu- rification was obtained by chromatography on immobilized wheat germ agglutinin. Specific saxitoxin-binding activity of the purified material averaged 1500 pmol of saxitoxin bound per mg of protein. Three bands were present in this material on sodium dodecyl sulfate/polyacrylamide gel electrophoresis. The purified material sedimented on a sucrose gradient with an