STRUCTURAL DIFFERENCES BETWEEN TWO SPECIES OF ACETYLCHOLINESTERASE FROM Torpedo calif. ELECTRIC ORGAN. Hai Won Chang, Philip Barnett*, Ernest Bock* and Terrone L. Rosenberry. Columbia University, 630 West 168 Street, New York, NY 10032 and Case Western Reserve University, Cleveland, OH 44106.
Acetylcholinesterase in electric organ of Torpedo californica consists of two classes: a collagen containing 17S form, extractable by high ionic strength buffers with antiproteolytic agents and a hydrophobic 6S form, extractable by detergent solution. These two forms of the enzyme were purified by affinity chromatography and a small amount of acetylcholine receptor contaminant was removed by adsorption to alpha-bungarotoxin-Sepharose 4B. Both the 17S and 6S forms contain free sulfhydryl groups, and treatment with 50mM N-ethylmaleimide or iodoacetamide produces slow inactivation with pseudo-first order kinetics. SDS polyacrylamide gel electrophoresis indicates that while the 6S enzyme is composed of only 66K catalytic subunits, the 17S enzyme consists of slightly larger 70 K MW catalytic subunits as well as two noncatalytic subunits of 103 K and 54 K MW. Amino acid analysis indicates that the 54 K subunit is collagen like. Comparison of the amino acid composition of the two catalytic subunits shows that the 66 K MW species is significantly enriched in amino acids with aliphatic side chains. whereas the 70 K species is higher in polar amino acids, suggesting that the catalytic subunits themselves are responsible for the difference in the solubility of the two forms of the enzyme. (Supported by grants from NSF #PCM-8022832 and the Muscular Dystrophy Association of America.)