Polypeptide composition of the alpha-latrotoxin receptor. High affinity binding protein consists of a family of related high molecular weight polypeptides complexed to a low molecular weight protein.
alpha-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 alpha-latrotoxin, we have studied alpha-latrotoxin-binding proteins from bovine and rat brain. Proteins purified by affinity chromatography on immobilized alpha-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 alpha-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 alpha-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 Südhof, T. C. (1991) Nature 353, 65-68). Surprisingly, the M(r) 79,000 and 43,000 proteins were also found in tissues insensitive to alpha-latrotoxin action. Since these proteins do not bind 125I-alpha-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 alpha-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. alpha-Latrotoxin binding correlated with the presence of the M(r) 160,000-220,000 proteins and M(r) 29,000 polypeptide, and alpha-latrotoxin formed stable complexes with the M(r) 160,000-220,000 proteins. Accordingly, the alpha-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.
Petrenko
A G
AG
Howard Hughes Medical Institute, University of Texas Southwestern Medical School, Dallas 75235.
Lazaryeva
V D
VD
Geppert
M
M
Tarasyuk
T A
TA
Moomaw
C
C
Khokhlatchev
A V
AV
Ushkaryov
Y A
YA
Slaughter
C
C
Nasimov
I V
IV
Südhof
T C
TC
D000595:Amino Acid Sequence; D000818:Animals; D001923:Brain Chemistry; D002135:Calcium-Binding Proteins; D002417:Cattle; D002462:Cell Membrane / Q000737:chemistry; D002499:Centrifugation, Density Gradient; D004492:Edetic Acid / Q000494:pharmacology; D008562:Membrane Glycoproteins / Q000737:chemistry; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009419:Nerve Tissue Proteins / Q000737:chemistry; D010446:Peptide Fragments / Q000737:chemistry; D010455:Peptides / Q000032:analysis* / Q000737:chemistry; D051381:Rats; D011950:Receptors, Cholinergic / Q000032:analysis* / Q000737:chemistry; D018000:Receptors, Peptide; D013111:Spider Venoms / Q000378:metabolism; D050857:Synaptotagmins
D002135:Calcium-Binding Proteins; D008562:Membrane Glycoproteins; D009419:Nerve Tissue Proteins; D010446:Peptide Fragments; D010455:Peptides; D011950:Receptors, Cholinergic; D018000:Receptors, Peptide; D013111:Spider Venoms; C034800:alpha-latrotoxin receptor; D050857:Synaptotagmins; C020763:alpha-latrotoxin; D004492:Edetic Acid