人工解析提高串联质谱鉴定肽段的数据利用度

Improving data utilization of tandem time-of flight mass spectrometry for peptides identification by manual interpretation

为降低蛋白质鉴定的假阴性,利用标准蛋白和实际样品,探讨影响基质辅助激光解吸电离–串联飞行时间质谱数据利用度的原因.发现误差设置偏小、翻译后修饰、异常酶解和氨基酸突变等是导致蛋白质数据库检索不匹配目标肽段的主要原因.实验表明,典型情况下一级和二级质谱误差可设定在0.3Da.对未与数据库匹配的串联质谱进行亚胺离子辅助的人工解析发现,色氨酸双氧化、丝氨酸乙酰胺化和半胱氨酸丙酰胺化等不常见修饰,酪氨酸与苯丙氨酸、天冬氨酸与谷氨酸两两之间的突变和非特异性酶解等现象,该研究可提高质谱鉴定的数据利用度.

To reduce the false negative for protein identification based on mass spectrometry （MS）, standard pro-teins and practical samples were used to explore the influence of the data utilization on matrix-assisted laser desorp-tion ionization-tandem time-of-flight mass spectrometry. It tion, irregular digestion and gene mutation were the main was found that mass tolerance, post-translational modifica- factors leading to the mismatch between protein database and targeted peptides. After manual explanation assisted by immonium ions in low m/z region, the mass error of first order MS and second order MS could be set to 0.3 Da typically. In addition, the dioxidation of tryptophan, propionamidation of cysteine, carbamidomethy serine and the methionine to lose 48 Da was detected as an uncommon modification. Furthermore, the mutation between tyrosine and phenylalanine and the mutation between aspartate and glutamate were also found. All the above factors can lead to the failure of matching tandem mass data by Mascot da-tabase searching. Results show that manual analysis of the ＂unused＂ MS/MS data will improve the protein and pep-tide identification utilization effectively.