Purification and characterization of iotrochotin, a novel toxin from the Caribbean sponge Iotrochota birotulata, which selectively permeabilizes synaptosomes.
A protein termed iotrochotin (IOT) was isolated from the exudate of the Caribbean sponge Iotrochota birotulata using as an assay its stimulation of the release of radioactivity from synaptosomes preloaded with [3H]choline. Sephadex G-50 chromatography of the exudate produced one peak, with a mol. wt of approximately 18,000, which was further resolved into two active fractions by anion exchange chromatography. The more tightly bound of the two fractions was characterized and referred to as IOT. The action of IOT was essentially complete by 0.5-1.0 min and was independent of the Ca2+ or Na+ content of the incubation mixture. Released radioactivity included about 50% each of [3H]acetylcholine and [3H]choline. Release of radioactivity increased as a function of IOT concentration and then reached a maximum. Extrapolated asymptotic release was nearly equal to that obtained by lysing the synaptosomes. IOT also released radioactivity from synaptosomes which had been preincubated with other tritiated neurotransmitters or with 2-[3H]deoxy-D-glucose. Lactate dehydrogenase and choline acetyltransferase activities were not released from synaptosomes by treatment with IOT, but were released by digitonin. IOT therefore releases some of the smaller molecular weight components of synaptosomes, but does not permeabilize the synaptosomal membrane in the same way as digitonin treatment.
Martin
J V
JV
Department of Biology, Rutgers University, Camden, NJ 08102.
Koenig
M L
ML
McClure
W O
WO
D000818:Animals; D002794:Choline / Q000378:metabolism; D002845:Chromatography; D066298:In Vitro Techniques; D007700:Kinetics; D008297:Male; D008387:Marine Toxins / Q000302:isolation & purification* / Q000633:toxicity; D008970:Molecular Weight; D018377:Neurotransmitter Agents / Q000378:metabolism; D010539:Permeability; D011161:Porifera; D051381:Rats; D017207:Rats, Sprague-Dawley; D013574:Synaptosomes / Q000187:drug effects* / Q000378:metabolism
D008387:Marine Toxins; D018377:Neurotransmitter Agents; C077893:iotrochotin; D002794:Choline