食品科学

• 生物工程 • 上一篇    下一篇

凡纳滨对虾过敏原精氨酸激酶的质谱鉴定及其免疫交叉反应研究

孙一帆,黄建芳,向军俭*,王彩霞,陈成枫   

  1. 暨南大学 广东省分子免疫与抗体工程重点实验室,广东 广州 510632
  • 出版日期:2015-01-15 发布日期:2015-01-16

Proteomics and Immunological Analysis of Arginine Kinase, an Important Shrimp Allergen from Litopenaeus vannamei

SUN Yifan, HUANG Jianfang, XIANG Junjian*, WANG Caixia, CHEN Chengfeng   

  1. Guangdong Province Key Laboratory of Molecular Immunology and Antibody Engineering, Jinan University, Guangzhou 510632, China
  • Online:2015-01-15 Published:2015-01-16

摘要:

目的:鉴定凡纳滨对虾分子质量40 kD过敏原,并分析其在有壳水产食物中的免疫交叉反应。方法:运用基质辅助激光解析串联飞行时间质谱仪(matrix assisted laser desorption ionization/time of flight mass spectrometry,MALDI-TOF/TOF-MS)鉴定凡纳滨对虾40 kD过敏原组分;使用软件BLAST、Clustal X2、MEGA 5.0分析该蛋白氨基酸序列的种间同源性;制备抗凡纳滨对虾过敏原精氨酸激酶的多克隆抗体,并与17 种常见有壳水生动物粗提液进行Western blotting,以分析该过敏原的免疫交叉反应。结果:凡纳滨对虾40 kD过敏原为精氨酸激酶(arginine kinase,AK);其氨基酸序列同源性分析显示AK在虾类(96%~100%)、蟹类(91%~93%)、贝类(49%~52%)、蟑螂(83%)中具有很高的同源性;Western blotting结果显示抗AK多抗与17 种不同虾类、蟹类、贝类的AK均能发生反应。结论:精氨酸激酶在甲壳类动物、软体动物、甚至昆虫中具有高度保守性,且能引起强免疫交叉反应,是有壳水生动物中的一种泛过敏原。

关键词: 凡纳滨对虾, 过敏原, 精氨酸激酶, 质谱, 序列同源性, 免疫交叉反应

Abstract:

This study aimed to identify the 40-kD allergen of Litopenaeus vannamei and to analyze its immune crossreactivity
among shellfish species. Several proteomics and immunological experiments were performed. By using matrixassisted
laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF/TOF-MS), the 40 kD allergen from
Litopenaeus vannamei was identified as arginine kinase (AK). Its amino acid sequence was compared with that of known
proteins by BLAST, Clustal X2 and MEGA 5. The results revealed that AK had high homology among shrimp (96%-100%),
crab (91%-93%), cockroach (83%) and mollusca (49%-52%).The purified AK was injected subcutaneously into mice
to produce specific polyclonal antibodies for analyzing its immune cross-reactivity by Western blotting. It was shown that
the proteins with relative molecular mass (Mr) of about 40 kD corresponding to AKs from 17 species of shellfish could be
recognized by the AK-specific polyclonal sera. The results of both proteomics and immunological analysis have shown that
AK is a pan-allergen among crustacean and mollusca.

Key words: Litopenaeus vannamei, allergen, arginine kinase, mass spectrometry, sequence homology, immune cross-reactivity

中图分类号: