Titulaer Mark K, Mustafa Dana A N, Siccama Ivar, Konijnenburg Marco, Burgers Peter C, Andeweg Arno C, Smitt Peter A E Sillevis, Kros Johan M, Luider Theo M
Department of Neurology, Laboratory of Neuro-Oncology, Clinical and Cancer Proteomics, Erasmus Medical Center, Dr, Molewaterplein 50, PO Box 2040, 3000 CA Rotterdam, The Netherlands.
BMC Bioinformatics. 2008 Mar 1;9:133. doi: 10.1186/1471-2105-9-133.
A Java application is presented, which compares large numbers (n > 100) of raw FTICR mass spectra from patients and controls. Two peptide profile matrices can be produced simultaneously, one with occurrences of peptide masses in samples and another with the intensity of common peak masses in all the measured samples, using the peak- and background intensities of the raw data. In latter way, more significantly differentially expressed peptides are found between groups than just using the presence or absence in samples of common peak masses. The software application is tested by searching angiogenesis related proteins in glioma by comparing laser capture micro dissected- and enzymatic by trypsin digested tissue sections.
By hierarchical clustering of the presence-absence matrix, it appears that proteins, such as hemoglobin alpha and delta subunit, fibrinogen beta and gamma chain precursor, tubulin specific chaperone A, epidermal fatty acid binding protein, neutrophil gelatinase-associated lipocalin precursor, peptidyl tRNA hydrolase 2 mitochondrial precursor, placenta specific growth hormone, and zinc finger CCHC domain containing protein 13 are significantly different expressed in glioma vessels. The up-regulated proteins in the glioma vessels with respect to the normal vessels determined by the Wilcoxon-Mann-Whitney test on the intensity matrix are vimentin, glial fibrillary acidic protein, serum albumin precursor, annexin A5, alpha cardiac and beta actin, type I cytoskeletal 10 keratin, calcium binding protein p22, and desmin. Peptide masses of calcium binding protein p22, Cdc42 effector protein 3, fibronectin precursor, and myosin-9 are exclusively present in glioma vessels. Some peptide fragments of non-muscular myosin-9 at the C-terminus are strongly up-regulated in the glioma vessels with respect to the normal vessels.
The less rigorous than in general used commercial propriety software de-isotope algorithm results in more mono-isotopic peptide masses and consequently more proteins. Centroiding of peptide masses takes place by taking the average over more spectra in the profile matrix. Cytoskeleton proteins and proteins involved in the calcium signaling pathway seem to be most up-regulated in glioma vessels. The finding that peptides at the C-terminus of myosin-9 are up-regulated could be ascribed to splicing or fragmentation by proteases.
本文介绍了一个Java应用程序，它可比较大量（n > 100）来自患者和对照的原始傅里叶变换离子回旋共振质谱。利用原始数据的峰强度和背景强度，可同时生成两个肽谱矩阵，一个显示肽质量在样本中的出现情况，另一个显示所有测量样本中共同峰质量的强度。通过这种方式，与仅使用共同峰质量在样本中的存在与否相比，在组间发现了更多显著差异表达的肽。通过比较激光捕获显微切割和胰蛋白酶消化的组织切片，在胶质瘤中搜索血管生成相关蛋白对该软件应用进行了测试。
通过对存在-缺失矩阵进行层次聚类，发现血红蛋白α和δ亚基、纤维蛋白原β和γ链前体、微管蛋白特异性伴侣A、表皮脂肪酸结合蛋白、中性粒细胞明胶酶相关脂质运载蛋白前体、肽基tRNA水解酶2线粒体前体、胎盘特异性生长激素以及含锌指CCHC结构域蛋白13等蛋白质在胶质瘤血管中表达存在显著差异。通过对强度矩阵进行威尔科克森-曼-惠特尼检验确定，相对于正常血管，胶质瘤血管中上调的蛋白质有波形蛋白、胶质纤维酸性蛋白、血清白蛋白前体、膜联蛋白A5、α心肌肌动蛋白和β肌动蛋白、I型细胞骨架角蛋白10、钙结合蛋白p22以及结蛋白。钙结合蛋白p22、Cdc42效应蛋白3、纤连蛋白前体和肌球蛋白9的肽质量仅存在于胶质瘤血管中。相对于正常血管，胶质瘤血管中C端非肌肉肌球蛋白9的一些肽片段强烈上调。
与一般使用的商业专有软件去同位素算法相比，该算法不太严格，导致更多的单同位素肽质量，从而发现更多的蛋白质。肽质量的峰位确定是通过对谱矩阵中更多光谱取平均值来实现的。细胞骨架蛋白和参与钙信号通路的蛋白在胶质瘤血管中似乎上调最为明显。肌球蛋白9 C端肽上调这一发现可能归因于蛋白酶的剪接或切割。
