【摘要】 目的:通过模拟肠胃液消化试验,分析锯缘青蟹肌肉中过敏原(原肌球蛋白)的消化特性以及消化对过敏蛋白致敏性的影响。方法:采用美国药典提供的模拟胃、肠液配方,测定青蟹原肌球蛋白和肌原纤维蛋白在体外模拟肠胃环境中的消化稳定性。以十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)和免疫印迹法(Western-blot)进行分析。结果:在模拟胃液反应中,非过敏原蛋白尤其是肌球蛋白重链和肌动蛋白等被胃蛋白酶快速降解,而原肌球蛋白在60min时仍未被完全分解。在模拟肠液反应中,相对于致敏蛋白,肌球蛋白重链、肌动蛋白等非过敏蛋白在较短的时间内被分解,原肌球蛋白尽管120min后几乎被完全分解,但会产生一些较稳定的蛋白降解片段。以免疫印迹法检测过敏患者血清,结果分子质量为34kDa的原肌球蛋白降解产物仍具有过敏原性。结论:蟹类过敏蛋白相对于非过敏蛋白具有较高的消化稳定性,而且其高分子质量降解产物仍可能引起过敏反应。更多还原

【Abstract】 Objective:To analyze the digestive stability of a major crustacean allergen(tropomyosin)from the muscle of mud crab(Scylla serrata) and the effect of gastrointestinal digestion on the allergenicity of tropomyosin in vitro.Methods:The composition of standard simulated gastric fluid(SGF) and simulated intestinal fluid(SIF) was based on the United States Pharmacopeia.The stability of allergenic protein tropomyosin and myofibrillar proteins from the muscle of mud crab to SGF and SIF was tested in vitro, respectively.The digestion was evaluated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis(SDS-PAGE) and Western blot.Results:In SGF assay, nonallergenic proteins especially myosin heavy chain(MHC) and actin were rapidly degraded within a short period of time, while tropomyosin could not be completely degraded within 60 min.In SIF assay, compared with tropomyosin, nonallergenic proteins such as MHC and actin were degraded within short time intervals.Though the original tropomyosin band was almost completely degraded within 120 min, some stable fragments could still be identified.Western blot analysis using human serum that had clinical history of crab allergy indicated that a degraded fragment with size of 34 kDa is still IgE-reactive.Conclusion:our present results indicated that crab tropomyosin is more resistant to proteinase digestion than non-allergenic proteins and the higher molecular mass fragment from tropomyosin after enzymatic treatment is still possible to cause hypersensitivity.更多还原