液相色谱-线性离子阱串联质谱法检测人正常晶状体水溶性蛋白质

Normal water-soluble proteins detection of human lens by liquid chromatography-linear ion trap tandem mass spectrometry

目的检测和分析人正常晶状体水溶性蛋白质的组成和功能。方法通过聚丙烯酰胺凝胶和液相色谱分离晶状体中的水溶性蛋白质,应用线性离子阱串联质谱仪鉴定蛋白质。运用基因本体论(geneontology,GO)对鉴定出的蛋白质的表达功能进行富集和缺失分析。结果鉴定出人正常水溶性晶状体蛋白质242个,包括许多酶类等重要的功能性蛋白。鉴定出的晶状体水溶性蛋白质参与细胞组成显示富集结果有:眼晶状体结构组成、细胞质、细胞内、细胞内部件、细胞骨架;参与分子功能显示富集结果有:催化活性、结构分子活性、氧化还原酶活性、异构酶活性;参与生物过程显示富集结果有:糖酵解、戊糖磷酸旁路、细胞大分子分解代谢过程、NADP代谢过程、烟酰胺代谢过程、吡啶核苷酸代谢过程、NADPH再生、细胞成分结构和生物合成的调节、以微管为基础的过程。运用GO分析结果表明,鉴定出蛋白质的功能与晶状体正常的生理活动密切相关,进一步证实鉴定结果的可靠性。结论线性离子阱串联质谱仪在晶状体水溶性蛋白质鉴定方面显示独特的优势。采用基于GO的功能分析等生物信息学分析方法是对蛋白质组学研究的有益补充。

Objective To detect and analyze the compositions and functions of water-soluble proteins of normal human lens.Methods The water-soluble proteins of normal human lens samples were separated by polyacrylamide gels and liquid chromatography.Linear ion trap tandem mass spectrometry was used for proteins identification.The function analyses of identified proteins were based on over-represented and under-represented annotations of gene ontology（GO）.Results Two hundreds and forty-two water-soluble proteins of normal human lens were identified,including various enzymes and other important functional proteins.Overrepresented annotations of cellular component of identified lens proteins included structural constituent of eye lens,cytoplasm,intracellular,intracellular part,cytoskeleton;Overrepresented annotations of molecular function included catalytic activity,structural molecule activity,oxidoreductase activity,isomerase activity;Overrepresented annotations of biological process included glycolysis,pentose-phosphate shunt,cellular macromolecule catabolic process,NADP metabolic process,nicotinamide metabolic process,pyridine nucleotide metabolic process,NADPH regeneration,regulation of cellular component organization and biogenesis,microtubule-based process.Analytic result of GO showed the functions of identified proteins were closely related to physical activity of normal lens,and further confirmed the reliability of proteins identification.Conclusions The linear ion trap tandem mass spectrometry has unique advantages in water-soluble proteins identification of normal human lens.Bioinformatic tools such as GO-based functional analyses are beneficial for proteomics studies.