Choudhary Gargi, Wu Shiaw-Lin, Shieh Paul, Hancock William S
Thermo Finnigan, San Jose, California 95134, USA.
J Proteome Res. 2003 Jan-Feb;2(1):59-67. doi: 10.1021/pr025557n.
This study uses multiple enzyme digests to increase the sequence coverage of proteins identified by the shotgun sequencing approach to proteomic analysis. The enzymes used were trypsin, Lys-C, and Asp-N, which cleave at arginine and lysine residues, lysine, and aspartic acid residues, respectively. This approach was evaluated with the glycoprotein, tissue plasminogen activator, t-PA and gave enhanced sequence coverage, compared with a single enzymatic digest. The approach was then evaluated with a complex proteomic sample, namely plasma. It was found that trypsin and Lys-C were able to detect overlapping but distinct sets of proteins and a digital recombination of the data gave a significant increase in both the number of protein identifications as well as an increase in the number of peptides identified per protein (which improves the certainty of the assignment).
本研究使用多种酶切方法来增加通过鸟枪法蛋白质组学分析鉴定的蛋白质的序列覆盖率。所使用的酶分别是胰蛋白酶、Lys-C和天冬酰胺酶N，它们分别在精氨酸和赖氨酸残基、赖氨酸以及天冬氨酸残基处切割。与单一酶切相比，该方法在糖蛋白组织型纤溶酶原激活剂（t-PA）上进行了评估，并获得了更高的序列覆盖率。然后，该方法在一个复杂的蛋白质组学样本即血浆上进行了评估。结果发现，胰蛋白酶和Lys-C能够检测到重叠但不同的蛋白质组，数据的数字重组显著增加了蛋白质鉴定的数量以及每个蛋白质鉴定出的肽段数量（这提高了鉴定的确定性）。
