Kline Kelli G, Frewen Barbara, Bristow Michael R, Maccoss Michael J, Wu Christine C
Department of Pharmacology, University of Colorado School of Medicine, Aurora, Colorado 80045, USA.
J Proteome Res. 2008 Nov;7(11):5055-61. doi: 10.1021/pr800239e. Epub 2008 Sep 20.
Proteomics research is beginning to expand beyond the more traditional shotgun analysis of protein mixtures to include targeted analyses of specific proteins using mass spectrometry. Integral to the development of a robust assay based on targeted mass spectrometry is prior knowledge of which peptides provide an accurate and sensitive proxy of the originating gene product (i.e., proteotypic peptides). To develop a catalog of "proteotypic peptides" in human heart, TRIzol extracts of left-ventricular tissue from nonfailing and failing human heart explants were optimized for shotgun proteomic analysis using Multidimensional Protein Identification Technology (MudPIT). Ten replicate MudPIT analyses were performed on each tissue sample and resulted in the identification of 30 605 unique peptides with a q-value < or = 0.01, corresponding to 7138 unique human heart proteins. Experimental observation frequencies were assessed and used to select over 4476 proteotypic peptides for 2558 heart proteins. This human cardiac data set can serve as a public reference to guide the selection of proteotypic peptides for future targeted mass spectrometry experiments monitoring potential protein biomarkers of human heart diseases.
蛋白质组学研究正开始从更为传统的蛋白质混合物鸟枪法分析扩展到包括使用质谱对特定蛋白质进行靶向分析。基于靶向质谱开发可靠检测方法的一个不可或缺的前提是,要事先知道哪些肽段能够准确且灵敏地代表原始基因产物(即蛋白质型肽段)。为了建立人类心脏中的“蛋白质型肽段”目录,对来自非衰竭和衰竭人类心脏外植体左心室组织的TRIzol提取物进行了优化,以便使用多维蛋白质鉴定技术(MudPIT)进行鸟枪法蛋白质组分析。对每个组织样本进行了十次重复的MudPIT分析,结果鉴定出30605个q值≤0.01的独特肽段,对应于7138种独特的人类心脏蛋白质。评估了实验观察频率,并用于为2558种心脏蛋白质选择了4476多个蛋白质型肽段。这个人类心脏数据集可作为公共参考,为未来监测人类心脏病潜在蛋白质生物标志物的靶向质谱实验选择蛋白质型肽段提供指导。