应用iTRAQ定量蛋白质组学技术筛选无乳链球菌鱼源株与牛源株差异表达蛋白

Quantitative identification of differential proteins in Streptococcus agalactiae piscine strain and bovine strain using i TRAQ

为揭示不同宿主来源无乳链球菌致病机制,提取无乳链球菌鱼源株GD201008-001和牛源株ATCC13813的全菌蛋白质,经iTRAQ试剂标记后进行质谱鉴定,质谱数据用软件Mascot2.2和Proteome Discoverer1.4进行查库(UniProt数据库)鉴定及定量分析,并利用blast2go软件对差异蛋白质进行GO(gene ontology)功能注释。共鉴定出在鱼源株GD201008-001和牛源株ATCC13813中差异表达的蛋白质17个(P<0.05),其中在ATCC13813中上调表达的蛋白质3个(比值>1.500),下调表达的蛋白质14个(比值<0.667)。GO分析预测结果表明这些蛋白质主要参与15个生物学功能,涉及代谢过程、细胞成分、结合、催化活性、结构分子活性等。其中cpsIVK在牛源株ATC13813中下调最显著,其功能为荚膜生物合成蛋白质,与无乳链球菌荚膜合成相关。进一步验证发现,鱼源株GD201008-001抵抗小鼠巨噬细胞RAW264.7吞噬的能力显著高于cpsIVK下调的牛源株ATCC13813(P<0.01),提示cpsIVK可能在鱼源无乳链球菌逃避宿主免疫细胞吞噬过程中发挥功能。

To identify differentially expressed proteins in Streptococcus agalactiae (S.galactiae) piscine strain (GD201008-001) and bovine strain (ATCC13813) by iTRAQ,and provide new clues for exploring the pathogenic mechanism of S.galactiaestrains isolated from different hosts,cellular proteins were extracted from cultures of S.galactiae GD201008-001 and ATCC13813,and labeled with isobaric tags for relative and absolute quantitation (i TRAQ).Differentially expressed proteins were identified with LC-MS/MS.The mass spectrometry data was analyzed by Mascot2.2 and Proteome Discoverer1.4 and subjected to biological information analysis by blast2 go.A total of 17 differentially expressed proteins were revealed in GD201008-001 and ATCC13813 (P < 0.05),among which 3 proteins were up-regulated (ratio > 1.5) and 14 were down-regulated (ratio < 0.667) in ATCC13813.These proteins are involed in 15 biological functions including metabolic process,cell composition,binding,catalytic activity,and hypothetical proteins.The most down-regulated protein cpsIVK encoding capsule biosynthesis protein was related to S.agalactiae capsular synthesis.Further experiments showed that the antiphagocytosis of GD201008-001 resisting murine macrophages RAW264.7 was significantly higher than ATCC13813 (P < 0.01) in which cpsIVK was down-regulated,suggesting that cpsIVK may play a role in the resistance to host immune cell phagocytosis of S.galactiae piscine strain.