~(60)Co-γ辐照对花生过敏原Ara h 2蛋白构象及致敏活性的影响
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摘要
为探讨辐照处理对花生Ara h 2蛋白结构与致敏活性的影响,采用不同剂量~(60)Co-γ辐照处理分离纯化所得到的花生过敏原Ara h 2蛋白,结合紫外扫描光谱、圆二色谱(CD)和聚丙烯酰胺凝胶电泳(SDS-PAGE)评估辐照处理后Ara h 2蛋白的结构变化,并用免疫印迹法和间接酶联免疫吸附法检测辐照处理后Ara h 2的抗原性变化。结果表明,~(60)Co-γ辐照处理可以显著改变花生Ara h 2蛋白的构象,使其降解、发生交联。随着辐照剂量的增大,Ara h 2蛋白与抗体的结合能力呈逐渐下降趋势,且与蛋白紫外吸光度的增强和α-螺旋含量的降低呈现良好的相关性。当辐照剂量为10 kGy时,可基本破坏Ara h 2蛋白的结构和免疫活性。~(60)Co-γ辐照处理可以有效降低花生过敏原Ara h 2蛋白的致敏性,这为花生脱敏技术的研究提供了新思路。
In order to investigate the effect of irradiation on the changes of structural and antigenic properties of Ara h 2, the separated peanut allergen Ara h 2 was irradiated by ~(60)Co-γ rays at different doses. The structures were characterized by ultraviolet absorption spectrum, circular dichroism(CD), and sodium dodecyl sulfate-polyacrylamide gel electrophoresis(SDS-PAGE), respectively. The antigenicity of the irradiated Ara h 2 was evaluated by Western blotting and indirect ELISA. Results showed that the conformation of Ara h 2 was significantly changed after ~(60)Co-γ irradiation treatment, which caused denaturation and aggregation of Ara h 2. The IgG binding capacity of Ara h 2 was reduced with the increasing irradiation doses, which correlated well with the rise of UV absorption intensity and a loss of α-helix content. Moreover, the structure and antigenicity of Ara h 2 were almost completely destroyed when the irradiation dose was up to 10 kGy. It indicated that ~(60)Co-γ irradiation can effectively reduce the allergenicity of Ara h 2, which may provide a new method on peanut desensitization technology.
In order to investigate the effect of irradiation on the changes of structural and antigenic properties of Ara h 2, the separated peanut allergen Ara h 2 was irradiated by ~(60)Co-γ rays at different doses. The structures were characterized by ultraviolet absorption spectrum, circular dichroism(CD), and sodium dodecyl sulfate-polyacrylamide gel electrophoresis(SDS-PAGE), respectively. The antigenicity of the irradiated Ara h 2 was evaluated by Western blotting and indirect ELISA. Results showed that the conformation of Ara h 2 was significantly changed after ~(60)Co-γ irradiation treatment, which caused denaturation and aggregation of Ara h 2. The IgG binding capacity of Ara h 2 was reduced with the increasing irradiation doses, which correlated well with the rise of UV absorption intensity and a loss of α-helix content. Moreover, the structure and antigenicity of Ara h 2 were almost completely destroyed when the irradiation dose was up to 10 kGy. It indicated that ~(60)Co-γ irradiation can effectively reduce the allergenicity of Ara h 2, which may provide a new method on peanut desensitization technology.
引文
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[27] 王梁燕,洪奇华,孙志明,华跃进.电子束辐照技术在生命科学中的应用[J].核农学报,2018,32(2):283-290
[28] 胡欣欣.γ射线照射对蛋白质结构和功能影响的研究[D].济南:山东大学,2015:10-29
[29] Luo C P,Hu C Q,Gao J Y,Li X,Wu Z H,Yang A S,Chen H B.A potential practical approach to reduce Ara h 6 allergenicity by gamma irradiation [J].Food Chemistry,2013,136(3):1141-1147
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[31] Meng X Y,Li X,Wang X K,Gao J Y,Yang H,Chen H B.Potential allergenicity response to structural modification of irradiated bovine α-lactalbumin[J].Food & Function,2016,7(7):3102-3110
[2] Zhou Y,Wang J S,Yang X J,Lin D H,Gao Y F,Su Y J,Yang S,Zhang Y J,Zheng J J.Peanut allergy,allergen Composition,and methods of reducing allergenicity:A review[J].International Journal of Food Science,2013,2013:1-8
[3] Peters R L,Koplin J J,Gurrin L C,Dharmage S C,Wake M,Ponsonby A L,Tang M L K,Lowe A J,Matheson M,Dwyer T,Allen K J.The prevalence of food allergy and other allergic diseases in early childhood in a population-based study:Health Nuts age 4-year follow-up[J].Journal of Allergy and Clinical Immunology,2017,140(1):145-153
[4] Kotz D,Simpson C R,Sheikh A.Incidence,prevalence,and trends of general practitioner-recorded diagnosis of peanut allergy in England,2001 to 2005[J].Journal of Allergy and Clinical Immunology,2011,127(3):623-630
[5] Zhuang Y H,Dreskin S C.Redefining the major peanut allergens[J].Immunologic Research,2013,55(1/3):125-134
[6] Koppelman S J,Vlooswijk R A,Knippels L M,Hessing M,Knol E F,van Reijsen F C,Bruijnzeel-Koomen C A.Quantification of major peanut allergens Ara h 1 and Ara h 2 in the peanut varieties Runner,Spanish,Virginia,and Valencia,bred in different parts of the world [J].Allergy,2001,56(2):132-137
[7] 李欣,陈红兵.食物过敏原构象性表位鉴别的研究进展[J].食品科学,2012,33(17):279-283
[8] Barre A,Borges J P,Culerrier R,Rougé P.Homology modelling of the major peanut allergen Ara h 2 and surface mapping of IgE-binding epitopes[J].Immunology Letters,2005,100(2):153-158
[9] Mueller G A,Gosavi R A,Pomés A,Wünschmann S,Moon A F,London R E,Pedersen L C.Ara h 2:Crystal structure and IgE binding distinguish two subpopulations of peanut allergic patients by epitope diversity[J].Allergy,2011,66(7):878-885
[10] 徐晨,白婵,耿胜荣,熊光权,张同,鉏晓艳,饶丹华,廖涛.60Co-γ射线辐照对草鱼贮藏品质的影响[J].核农学报,2018,32(6):1116-1124
[11] 顾可飞,高美须,李春红,潘家荣.辐照降低食物致敏性的研究进展[J].核农学报,2006,20(6):524-526
[12] Vaz A F M,Souza M P,Vieira L D,Aguiar,J S,Silva,T G,Medeiros,P L.High doses of gamma radiation suppress allergic effect induced by food lectin[J].Radiation Physics and Chemistry,2013,85(4):218-226
[13] Gomaa A,Boye J.Impact of irradiation and thermal processing on the immunochemical detection of milk and egg allergens in foods[J].Food Research International,2015,74(10):275-283
[14] 李新,熊光权,廖涛,鉏晓艳,耿胜荣,程薇.小龙虾虾肉辐照后理化指标与蛋白质性质分析[J].核农学报,2016,30(10):1941-1949
[15] 王烁,张春红,李淑荣.辐照处理对花生致敏蛋白的影响[J].花生学报,2009,38(2):1-5
[16] 许舒婷,高美须,支玉香,潘家荣,李淑荣,王志东,刘超超,赵杰,兰丽平,徐迎阳.电子束辐照对花生过敏原免疫原性及生化性质影响的研究[J].核农学报,2012,26(7):1006-1011
[17] 胡纯秋.花生过敏原Ara h 2 的研究[D].南昌:南昌大学,2010:15-29
[18] 刘珂,熊丽姬,高金燕,陈红兵,佟平.加工对鸡蛋中四种主要过敏原结构的影响[J].食品科学,2017,38(23):51-61
[19] 丁晓岚,高红旗.圆二色光谱技术应用和实验方法[J].实验技术与管理,2008,25(10):48-52
[20] 王田虎.药物与生物分子相互作用的光谱特性分析与研究[D].南京:南京航空航天大学,2011:8-17
[21] 李艳青.蛋白质氧化对鲤鱼蛋白结构和功能性的影响及其控制技术[D].哈尔滨:东北农业大学,2013:7-24
[22] 黄汉昌,姜招峰,朱宏吉.紫外圆二色光谱预测蛋白质结构的研究方法[J].化学通报,2007(7):501-506
[23] 白雨鑫,李欣,赵丽霞,陈红兵,高金燕.食物过敏原结构变化的光谱学研究进展[J].食品科学,2012,33(17):284-289
[24] 沈星灿,梁宏,何锡文,王新省.圆二色光谱分析蛋白质构象的方法及研究进展[J].分析化学,2004,32(3):388-394
[25] Marsh J,Rigby N,Wellner K,Reese G,Knulst A,Akkerdaas J,van Ree R,Radauer C,Lovegrove A,Sancho A,Mills C,Vieths S,Hoffmann-Sommergruber K,Shewry P R.Purification and characterisation of a panel of peanut allergens suitable for use in allergy diagnosis[J].Molecular Nutrition & Food Research,2008,52(Sup 2):272-285
[26] Rahaman T,Vasiljevic T,Ramchandran L.Effect of processing on Conformational changes of food proteins related to allergenicity[J].Trends in Food Science & Technology,2016,49:24-34
[27] 王梁燕,洪奇华,孙志明,华跃进.电子束辐照技术在生命科学中的应用[J].核农学报,2018,32(2):283-290
[28] 胡欣欣.γ射线照射对蛋白质结构和功能影响的研究[D].济南:山东大学,2015:10-29
[29] Luo C P,Hu C Q,Gao J Y,Li X,Wu Z H,Yang A S,Chen H B.A potential practical approach to reduce Ara h 6 allergenicity by gamma irradiation [J].Food Chemistry,2013,136(3):1141-1147
[30] Byun M W,Lee J W,Yook H S,Jo C,Kim H Y.Application of gamma irradiation for inhibition of food allergy[J].Radiation Physics and Chemistry,2002,63(3/6):369-370
[31] Meng X Y,Li X,Wang X K,Gao J Y,Yang H,Chen H B.Potential allergenicity response to structural modification of irradiated bovine α-lactalbumin[J].Food & Function,2016,7(7):3102-3110