Polypeptide composition of the α-latrotoxin receptor. High affinity binding protein consists of a family of related high molecular weight polypeptides complexed to a low molecular weight protein

α-Latrotoxin is a vertebrate neurotoxin from black widow spider venom that causes massive neurotransmitter release. In order to gain insight into the mechanism of action of α-latrotoxin, we have studied α-latrotoxin- binding proteins from bovine and rat brain. Proteins purified by affinity chromatography on immobilized α-latrotoxin were investigated. Two sets of proteins were isolated: 1) three polypeptides of M(r) 79,000, 65,000, and 43,000 that were eluted from immobilized α-latrotoxin by increasing KCl concentrations in the presence of Ca²⁺, and 2) a family of related proteins ranging in molecular weight from 160,000 to 220,000 and a low molecular weight component of M(r) 29,000 that were eluted from immobilized α- latrotoxin only after removal of Ca²⁺. Amino acid sequences of these proteins demonstrated that all of these proteins represent novel proteins except for the M(r) 65,000 polypeptide, which is identical with synaptotagmin (Petrenko, A. G., Perin, M. S., Davletov, B. A., Ushkaryov, Y. A., Geppert, M., and Sudhof, T. C. (1991) Nature 353, 65-68). Surprisingly, the M(r) 79,000 and 43,000 proteins were also found in tissues insensitive to α- latrotoxin action. Since these proteins do not bind ¹²⁵I-α-latrotoxin with high affinity, their purification probably is not physiologically significant. On the other hand, the fractions containing the M(r) 160,000- 220,000 and 29,000 polypeptides bound α-latrotoxin with high affinity. Sucrose gradient centrifugations and anion exchange chromatography suggested that most of the M(r) 160,000-220,000 proteins were complexed with the M(r) 29,000 protein. α-Latrotoxin binding correlated with the presence of the M(r) 160,000-220,000 proteins and M(r) 29,000 polypeptide, and α-latrotoxin formed stable complexes with the M(r) 160,000-220,000 proteins. Accordingly, the α-latrotoxin receptor consists of a high molecular weight protein (M(r) 160,000-220,000) that is complexed with one or several copies of an M(r) 29,000 polypeptide. In addition, the receptor is found in a less tight association with synaptotagmin but not with other polypeptides.