基于串联质谱标签的蛋白质组学技术分析海水流化冰辅助保鲜对大黄鱼肌肉蛋白质的影响

Tandem mass tag-based proteomics technology analyzing the effects of seawater slurry ice-assisted preservation on muscle proteins of Larimichthys crocea

为研究大黄鱼(Larimichthys crocea)蛋白质在海水流化冰辅助保鲜期内的变化,采用串联质谱标签(tandem mass tag,TMT)技术定量标记新鲜(贮藏0 d)大黄鱼及不同海水流化冰贮藏时间(7、23 d)下大黄鱼的肌肉蛋白质,筛选出不同贮藏时间下的差异表达蛋白,并对差异蛋白进行基因本体(gene ontology,GO)、直系同源簇/真核生物直系同源簇(clusters of orthologous groups/clusters of eukaryotic orthologous groups,COG/KOG)与京都基因和基因组数据库(Kyoto Encyclopedia of Genes and Genomes,KEGG)分析。结果显示,在3个比较组(7 d vs 0 d、23 d vs 7 d和23 d vs 0 d)中鉴定到的差异蛋白总数分别为215、133、269个,其中表达上调的差异蛋白数分别为117、37、113个,表达下调的差异蛋白数分别为98、96、156个。同时,3个比较组的共有差异蛋白为44个。GO分析表明,差异蛋白主要参与细胞过程、生物调节、细胞和细胞内组分、结合功能和催化活性等方面;COG/KOG分析表明,差异蛋白的功能活性主要集中在细胞骨架、碳水化合物转运和代谢、信号转导机制以及翻译后修饰、蛋白质周转和伴侣蛋白等方面;KEGG分析表明,差异蛋白共参与38条信号通路,且在贮藏后期(23 d),紧密连接和致病性大肠埃希菌感染2条信号通路中差异蛋白的参与数目最多,它们可能是造成大黄鱼新鲜度急剧下降的主要信号通路。44个共有差异蛋白与大黄鱼新鲜度指标进行的相关性分析表明,脆性X智力低下综合征相关蛋白2、肌醇-1-单磷酸酶、Rho GDP解离抑制剂1、锚蛋白-1和甲基四氢叶酸合酶结构域蛋白可以作为新鲜度指示蛋白。本研究为揭示大黄鱼在海水流化冰贮藏期内的品质变化机制、大黄鱼新鲜度评价和保鲜工艺改进提供了依据。

In order to study the changes in muscle proteins of Larimichthys crocea during the seawater slurry ice-assisted preservation period,tandem mass tag(TMT)-based proteomics technology was used to quantitatively analyze the muscle proteins of fresh(stored for 0 d)L.crocea and the samples preserved by seawater slurry ice for different storage times(7 and 23 d).The differential proteins under different storage times were screened out,and they were analyzed by gene ontology(GO),clusters of orthologous groups/clusters of eukaryotic orthologous groups(COG/KOG),and Kyoto Encyclopedia of Genes and Genomes(KEGG).The results showed that the total number of differential proteins identified in the three comparison groups(7 d vs 0 d,23 d vs 7 d,and 23 d vs 0 d)was 215,133 and 269,respectively.The number of up-regulated differential proteins was 117,37 and 113,respectively,and the number of down-regulated differential proteins was 98,96 and 156,respectively.At the same time,a total of 44 common differential proteins were identified in the three comparison groups.GO analysis showed that the differential proteins were mainly involved in cellular process,biological regulation,cell and intracellular components,binding function,and catalytic activity.COG/KOG analysis showed that the functional activities of differential proteins were mainly concentrated in cytoskeleton,carbohydrate transport and metabolism,signal transduction mechanisms and posttranslational modification,protein turnover,and chaperones.KEGG analysis showed that 38 signaling pathways were involved and the number of differential proteins involved in the two signaling pathways of tight junction and pathogenic Escherichia coli infection was the highest at the late storage(23 d),which might be the main metabolic pathways causing the sharp decline in the freshness of L.crocea.The correlation analysis between 44 common differential proteins with the freshness indexes of L.crocea showed that five freshness indicator proteins were screened out,including fragile X mental retardation syndrome-related protein 2,inositol-1-monophosphatase,Rho GDP-dissociation inhibitor 1,ankyrin-1,and methyltetrahydrofolate synthase domain-containing protein.This study provides a basis for revealing the quality change mechanism,the freshness evaluation,and the preservation process improvement of L.crocea during the storage period with seawater slurry ice.