| Budget Amount *help |
¥1,900,000 (Direct Cost: ¥1,900,000)
Fiscal Year 1992: ¥900,000 (Direct Cost: ¥900,000)
Fiscal Year 1991: ¥1,000,000 (Direct Cost: ¥1,000,000)
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| Research Abstract |
Among spiny lobster fishermen in Miyazaki Prefecture, bronchial asthma due to occupational sensitization with the soft coral Denronephthytia nipponica occurred. We found at least three different allergens with different molecular sizes which caused severe allergic skin reactions in the patients. We purified two major allergens, Den n1 and Den n2, with high molecular weights from the soft coral by ultrafiltration, gel filtration on Sephacryl s-200, and FPLC on a Mono-Q column. Both allergens caused severe allergic reactions when checked by the intradermal injection to the patients at a dose of less than 1 ng protein. Den n1 was the glycoprotein having a molecular weight greater than two million, and contained 30% neutral sugar as glucose. In SDS-polyacrylamide gel electrophoresis (PAGE) it showed two proteins bands corresponding to the molecular weights of 66,000 and 60,000. Both subunits gave the similar amino acid compositions in amino acid analysis. Den n2 showed a single band corresponding to the molecular weight of 80,000 in SDS-PAGE. It gave two bands, 54,000 and 21,000, under reduced condition. After electro-blotting to the PVDF membrane, the N-terminal amino acid-sequence was dtermined as Asp-Asp-Ile-Asn-Arg-Tyr-Ala-Phe-Asp-Asn-Lys-Ile-Asn-Asp-Lys-Leu-Phe-Asn-His-Trp-Glu-Ser-.
In the lower molecular weight fraction as well as fat-soluble fraction we detected the presence of both acetylcholine agonists and antagonists positive to the Dragendorff reagent. The purification of these compounds is under progress.
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