Cao P, Moini M
Department of Chemistry and Biochemistry, University of Texas at Austin 78712, USA.
Rapid Commun Mass Spectrom. 1998;12(13):864-70. doi: 10.1002/(SICI)1097-0231(19980715)12:13<864::AID-RCM244>3.0.CO;2-G.
Capillary electrophoresis/electrospray ionization (CE/ESI) high mass accuracy time-of-flight mass spectrometry was used for the first time to characterize small proteins using peptide mapping. To identify small proteins, the intact proteins were first analyzed to obtain their average molecular weights with errors less than 1 Da. On-line capillary electrophoresis mass spectrometry of the tryptic digests of these small proteins was then performed to obtain the accurate molecular weights of the peptides with accuracies of approximately 10 ppm. Next, this information was used for the identification of the proteins using a protein database. It was found that high mass accuracy is an effective tool in reducing the list of most-likely proteins generated by the database. In addition, on-line collision-induced dissociation of the completely or partially resolved capillary electrophoresis peaks of the protein digests was used to unambiguously identify the sequences of these peptides. Each CE/ESI-MS analysis used only 5 nL of sample containing approximately 120 fmol of each peptide in protein digests. The results indicate that the combination of capillary electrophoresis and high resolution, high mass accuracy time-of-flight mass spectrometry is a viable option for the identification of small proteins using peptide mapping.
首次使用毛细管电泳/电喷雾电离（CE/ESI）高精度飞行时间质谱法，通过肽图谱分析来表征小蛋白质。为鉴定小蛋白质，首先对完整蛋白质进行分析，以获得误差小于1 Da的平均分子量。然后对这些小蛋白质的胰蛋白酶消化产物进行在线毛细管电泳质谱分析，以获得精度约为10 ppm的肽段精确分子量。接下来，利用这些信息通过蛋白质数据库鉴定蛋白质。结果发现，高精度质量是减少数据库生成的最可能蛋白质列表的有效工具。此外，对蛋白质消化产物完全或部分分离的毛细管电泳峰进行在线碰撞诱导解离，以明确鉴定这些肽段的序列。每次CE/ESI-MS分析仅使用5 nL样品，其中蛋白质消化产物中每种肽段约含120 fmol。结果表明，毛细管电泳与高分辨率、高精度飞行时间质谱联用是通过肽图谱分析鉴定小蛋白质的可行选择。
