Yang Ziping, Harris Lyndsay E, Palmer-Toy Darryl E, Hancock William S
Barnett Institute and Department of Chemistry and Chemical Biology, Northeastern University, Boston, MA 02115, USA.
Clin Chem. 2006 Oct;52(10):1897-905. doi: 10.1373/clinchem.2005.065862. Epub 2006 Aug 17.
Glycoproteins are often associated with cancer and are important in serum studies, for which glycosylation is a common posttranslational modification.
We used multilectin affinity chromatography (M-LAC) to isolate glycoproteins from the sera of breast cancer patients and controls. The proteins were identified by HPLC-tandem mass spectrometry (MS/MS) analysis of the corresponding tryptic digests. We used the FuncAssociate Gene Ontology program for association analysis of the identified proteins. Biomarker candidates in these groups were comparatively quantitated by use of peak area measurements, with inclusion of an internal standard. We analyzed data for concordance within the ontology association groups for vector of change with the development of breast cancer.
Detection of the known low-concentration biomarker HER-2 (8-24 mug/L) enabled us to establish a dynamic range of 10(6), relative to the amount of albumin, for the depletion step. We then used ELISA to confirm this range. Proteins associated with lipid transport and metabolism, cell growth and maintenance, ion homeostasis, and protease inhibition were found to be differentially regulated in serum from women with breast cancer compared with serum from women without breast cancer.
M-LAC for isolation of the serum glycoproteome, coupled with liquid chromatography-MS/MS and the use of gene ontology associations, can be used to characterize large panels of candidate markers, which can then be evaluated in a particular patient population.
糖蛋白常与癌症相关，在血清研究中很重要，糖基化是一种常见的翻译后修饰。
我们使用多凝集素亲和色谱法（M-LAC）从乳腺癌患者和对照者的血清中分离糖蛋白。通过对相应胰蛋白酶消化产物的高效液相色谱-串联质谱（MS/MS）分析来鉴定蛋白质。我们使用FuncAssociate基因本体程序对鉴定出的蛋白质进行关联分析。通过峰面积测量并加入内标对这些组中的生物标志物候选物进行相对定量。我们分析了本体关联组内关于乳腺癌发展过程中变化向量的一致性数据。
检测已知的低浓度生物标志物HER-2（8 - 24微克/升）使我们能够确定相对于白蛋白量而言，去除步骤的动态范围为10^6。然后我们使用酶联免疫吸附测定法（ELISA）来确认这个范围。与脂质转运和代谢、细胞生长与维持、离子稳态以及蛋白酶抑制相关的蛋白质在乳腺癌女性血清中与无乳腺癌女性血清相比存在差异调节。
用于分离血清糖蛋白组的M-LAC，结合液相色谱-串联质谱以及基因本体关联的使用，可用于表征大量候选标志物，然后可在特定患者群体中进行评估。
