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Characterization of a domoic acid binding site from Pacific razor clam
In: Aquatic Toxicology. Elsevier Science: Tokyo; New York; London; Amsterdam. ISSN 0166-445X; e-ISSN 1879-1514
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| Trefwoorden |
Pseudo-nitzschia H. Peragallo in H. Peragallo & M. Peragallo, 1900 [WoRMS]; Siliqua patula (G. Dixon, 1789) [WoRMS]
Marien/Kust |
| Author keywords |
domoic acid; kainic acid; razor clam; glutamate receptor; kamate bindingprotein; receptor binding |
| Abstract |
The Pacific razor clam, Siliqua patula, is known to retain domoic acid, a water-soluble glutamate receptor agonist produced by diatoms of the genus Pseudo-nitzschia. The mechanism by which razor clams tolerate high levels of the toxin, domoic acid, in their tissues while still retaining normal nerve function is unknown. In our study, a domoic acid binding site was solubilized from razor clam siphon using a combination of Triton X-100 and digitonin. In a Scatchard analysis using [3H]kainic acid, the partially-purified membrane showed two distinct receptor sites, a high affinity, low capacity site with a KD (mean ± S.E.) of 28 ± 9.4 nM and a maximal binding capacity of 12 ± 3.8 pmol/mg protein and a low affinity, high capacity site with a mM affinity for radiolabeled kainic acid, the latter site which was lost upon solubilization. Competition experiments showed that the rank order potency for competitive ligands in displacing [3H]kainate binding from the membrane-bound receptors was quisqualate > ibotenate > IODOWILLARDIINE = AMPA = fluorowillardiine > domoate > kainate > -glutamate. At high micromolar concentrations, NBQX, NMDA and ATPA showed little or no ability to displace [3H]kainate. In contrast, Scatchard analysis using [3H]glutamate showed linearity, indicating the presence of a single binding site with a KD and Bmax of 500 ± 50 nM and 14 ± 0.8 pmol/mg protein, respectively. These results suggest that razor clam siphon contains both a high and low affinity receptor site for kainic acid and may contain more than one subtype of glutamate receptor, thereby allowing the clam to function normally in a marine environment that often contains high concentrations of domoic acid. |
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