CAJ | 학술논문

以13份甲壳类动物过敏患者血清的Western—blotting分析，确定克氏原螯虾主要过敏原为36ku蛋白质．通过盐析、等电点沉淀及热处理等方法纯化该蛋白质，以兔抗拟穴青蟹原肌球蛋白（Tropo-myosin，TM）多克隆抗体的Western—blotting分析，确定该蛋白质为TM．同源性分析表明，克氏原螯虾TM与南美白对虾TM、拟穴青蟹的氨基酸序列相似性较高（〉90％）．酶切位点预测显示，它分别有49个胰蛋白酶和6个胰凝乳蛋白酶的酶切位点．模拟胃肠液消化实验结果显示，纯化TM不易被胃蛋白酶降解，易于被胰蛋白酶和胰凝乳蛋白酶降解，进一步的Western—blotting和抑制性ELISA分析结果显示，其消化产物仍具有一定的免疫活性．采用蒸煮处理可降低TM的消化稳定性及免疫活性，且蒸煮处理时间越长，效果越显著．说明。TM为克氏原螯虾主要过敏原．与其他甲壳娄动物TM的序列相似件较高．
이13빈갑각류동물과민환자혈청적Western—blotting분석，학정극씨원오하주요과민원위36ku단백질．통과염석、등전점침정급열처리등방법순화해단백질，이토항의혈청해원기구단백（Tropo-myosin，TM）다극륭항체적Western—blotting분석，학정해단백질위TM．동원성분석표명，극씨원오하TM여남미백대하TM、의혈청해적안기산서렬상사성교고（〉90％）．매절위점예측현시，타분별유49개이단백매화6개이응유단백매적매절위점．모의위장액소화실험결과현시，순화TM불역피위단백매강해，역우피이단백매화이응유단백매강해，진일보적Western—blotting화억제성ELISA분석결과현시，기소화산물잉구유일정적면역활성．채용증자처리가강저TM적소화은정성급면역활성，차증자처리시간월장，효과월현저．설명。TM위극씨원오하주요과민원．여기타갑각루동물TM적서렬상사건교고．
The protein with molecular mass of 36 ku was detected from crustacean-allergic patients, suggesting it was the major allergen of by Western -blotting using 13 sera crayfish. The protein was further pu- rifled through the procedure of ammonium sulfate grading precipitation, isoelectric precipitation and thermal process. Purified protein was demonstrated to be TM by Western - blotting using polyclonal antibody against TM from Scylla paramamosain. Sequence analysis showed that crayfish TM shared high identities （〉 90 % ） with shrimp （Penaeus vannamei） and crab （Scylla paramamosain） . Cleavage site analysis results showed that crayfish TM contained 49 cleavage sites for trypsin and 6 cleavage sites for chymotrypsin. The simulated gastrointestinal fluid digestion showed that purified TM was more resistant to pepsin degradation, but more susceptible to trypsin and chymotrypsin degradation. Furthermore, the results of Western - blotting and inhi- bition ELISA showed that there was still a certain degree of immunoactivity in the digestion product. In addi- tion, the digestion stability and immunoactivity of TM were declined with the extension of cooking time. Thus, TM was the major allergen of crayfish, shared high identities with other crustacean TM.