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Bee venom neurotoxin (apamin): iodine labeling and characterization of binding sites.

1 
Apamin, the octadecapeptide from bee venom acting on the central nervous system, has been labeled with 125I at its single histidine residue. Apamin, monoiodoapamin, diiodoapamin, and iodide can be separated on thin-layer chromatography. Diiodoapamin retains about half the toxicity of the original peptide. Partial iodination with 125I, leading mainly to the monoiodo derivative, did not measurably decrease the toxicity. 
 
2 
Labeled apamin binds to the insoluble, but not to the soluble part of homogenates made from rat cerebral cortex. The order of binding to homogenates is rat forebrain > rostral part of the brain stem > cerebellum > caudal part of the brain stem > spinal cord. Grey substance of rabbit forebrain binds much better than white matter. The binding site is enriched within fractions of rat cerebral cortex which sediment between 10000 and 100000 × g. 
 
3 
Apamin also binds to peripheral organs. In rats, the sequence of specific apamin binding by different organs is cerebral cortex ≫ (20 ×) liver > kidney or ileum whereas binding by skeletal muscle, salivary glands, and testes is hardly measurable. In rabbits, the sequence is cerebral cortex ∼ liver > adrenals > cerebellum > kidney > medulla oblongata, whereas skeletal muscle and ileal mucosa do not bind to a measurable degree. Among all organs tested, bovine adrenal cortex binds the most. 
 
4 
Binding data with rat cerebral cortex were obtained under a variety of conditions. A single type of binding sites occurred with a density of about 1–3 pmol/g rat forebrain (wet weight). A very high affinity (Kd in the range of 10–25 pM) was found in both equilibrium and velocity measurements. 
 
5 
With increasing temperature apamin binding is diminished. Labeled apamin is slowly degraded by the supernatant of rat brain homogenate at 37°C and 25°C, but not at 5°C. 
 
 
 
 
 
High specificity, high affinity and prevalence of binding sites within the nervous system indicate, together with the rank order of potency of apamin derivatives, a role of binding in poisoning. However, final proof will need a direct comparison between binding and the action of apamin on isolated systems.