肺腺癌胸腔积液Shotgun蛋白质组学分析
|
摘要
目的获得人肺腺癌恶性胸腔积液详细的蛋白质组分信息,建立人类肺腺癌恶性胸腔积液蛋白质表达谱,以期为筛选出有潜力成为肺腺癌相关分子标志物的蛋白质奠定基础。
方法应用shotgun研究策略(单向凝胶电泳—液相色谱串联质谱)对5例病理明确诊断的肺腺癌病人胸腔积液的混合样品进行蛋白质组学研究,首先应用十二烷基磺酸钠—聚丙烯酰胺凝胶电泳(SDS-PAGE)初步分离胸腔积液蛋白样品,建立SDS凝胶电泳图谱,然后通过液相色谱串联质谱(1D-LC-LTQ系统,Finnigan线性离子阱串联质谱仪)进行分析,将质谱分析得到的数据应用SEQUEST软件及数据库搜索完成蛋白质组学鉴定,并对鉴定出的所有蛋白质从亚细胞定位、生物学途径、分子功能三个方面进行GOA分类。
结果应用shotgun蛋白质组学方式在人肺腺癌胸腔积液混合样品中成功鉴定出230种蛋白质,并使用GOA工具得到这些蛋白质亚细胞定位、生物学途径、分子功能三个方面的详细分类信息。在所有230种蛋白中,其中具有高可信度的蛋白质有123种(至少有两段独特肽序列与之匹配),可信度较低的蛋白质有107种(只有一段独特肽序列与之匹配)。在鉴定出的123种高可信度蛋白质之中,绝大多数蛋白质来源于血浆,只有7种蛋白质迄今为止尚无文献报道在血浆中出现,包括膜联蛋白A2、JUP蛋白、转化生长因子βig-h3蛋白、V-set及免疫球蛋白区域包含蛋白4、Ifapsoriasin2、肌动蛋白胞质型1(Actin,cytoplasmic 1)、Suprabasin。
结论通过shotgun蛋白质组学技术以及GOA注释,我们获得人肺腺癌恶性胸腔积液详细的蛋白质组份信息,初步建立了人肺腺癌恶性胸腔蛋白质表达谱。123种高可信度蛋白质的绝大多数来源于血浆,只有7种蛋白质迄今为止尚未见报道在血浆中出现,包括膜联蛋白A2、JUP蛋白、转化生长因子βig-h3蛋白、V-set及免疫球蛋白区域包含蛋白4、Ifapsoriasin2、肌动蛋白胞质型1(Actin,cytoplasmic1)、Suprabasin,这些蛋白质存在于恶性胸腔积液而不易在血浆中被检测出来,因此可能有潜力成为诊断肺腺癌的分子标志物,其中膜联蛋白A2因其具有调节细胞的粘附和生长增殖能力而更具特殊意义。
Objective To acquire protein profiles of malignant pleural effusion from lung adenocarcinoma patients, provide information on the composition of protein contents in human malignant pleural effusion, in order to detect the potential candidates of useful biomarkers with diagnostic value in the pleural effusion of lung adenocarcinoma.
Method Pleural effusion samples were collected from five lung adenocarcinoma patients, and a composite sample was analyzed by using shotgun strategy (one-dimensional gel electrophoresis liquid chromatography-tandem mass spectrometry,1D-LC-MS/MS). Pleural effusion samples were separated by means of Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE ) and commassie blue staining .Proteomic analysis was performed by LC-MS/MS using an LTQ linear IT mass spectrometer (Thermo. San Jose, CA, USA),then the proteins were identified by using SEQUEST software and protein database search .And we classified all of the proteins identified in this study according to Gene Ontology Annotation (GOA) in aspects of molecule function, Subcellular location, and biological process as well.
Results Applying the shotgun strategy, we identified in total 230 unique proteins from human pleural effusion of lung adenocarcinoma patients. We classified these 230 proteins identified in this study according to the GOA, all of them were performed functional classification, biological process classification and Subcellular location classification. 123 proteins were identified with higher confidence levels (at least two unique peptide sequences matched),while 107 other proteins were identified at the minimal confidence level (only one unique peptide sequence matched). Most of these 123 proteins that were identified with higher confidence levels have been reported in plasma , however, among these 123 proteins,7 proteins , JUP protein, Suprabasin,AnnexinA2,Transforming growth factor-beta-induced protein ig-h3 precursor ,V-set and immunoglobulin domain-containing protein 4 precursor , Ifapsoriasin 2 ,and Actin, cytoplasmic1 have not been reported in plasma ,and they may represent proteins specifically present in pleural effusion.
Conclusion By using shotgun proteomic approaches and gene ontologyannotation (GOA), we have got fundamental information on the composition of protein contents in human malignant pleural effusion and profiled the protein express of human pleural effusion from lung adenocarcinoma patients. Most of these 123 proteins identified with higher confidence levels from human pleural effusion of lung adenocarcinoma patients are originated from plasma. However, 7 proteins, such as JUP protein, Suprabasin, Annexin A2,Transforming growth factor-beta-induced protein ig-h.3 precursor ,V-set and immunoglobulin domain-containing protein 4 precursor , Ifapsoriasin 2 and Actin,cytoplasmic1have not been reported in plasma. These proteins could represent potential candidates of useful biomarkers that could not be readily detected in plasma but in malignant pleural effusion. AnnexinA2 is of special interest since it may play a role in the regulation of intercellular adhesion and cell proliferation.
方法应用shotgun研究策略(单向凝胶电泳—液相色谱串联质谱)对5例病理明确诊断的肺腺癌病人胸腔积液的混合样品进行蛋白质组学研究,首先应用十二烷基磺酸钠—聚丙烯酰胺凝胶电泳(SDS-PAGE)初步分离胸腔积液蛋白样品,建立SDS凝胶电泳图谱,然后通过液相色谱串联质谱(1D-LC-LTQ系统,Finnigan线性离子阱串联质谱仪)进行分析,将质谱分析得到的数据应用SEQUEST软件及数据库搜索完成蛋白质组学鉴定,并对鉴定出的所有蛋白质从亚细胞定位、生物学途径、分子功能三个方面进行GOA分类。
结果应用shotgun蛋白质组学方式在人肺腺癌胸腔积液混合样品中成功鉴定出230种蛋白质,并使用GOA工具得到这些蛋白质亚细胞定位、生物学途径、分子功能三个方面的详细分类信息。在所有230种蛋白中,其中具有高可信度的蛋白质有123种(至少有两段独特肽序列与之匹配),可信度较低的蛋白质有107种(只有一段独特肽序列与之匹配)。在鉴定出的123种高可信度蛋白质之中,绝大多数蛋白质来源于血浆,只有7种蛋白质迄今为止尚无文献报道在血浆中出现,包括膜联蛋白A2、JUP蛋白、转化生长因子βig-h3蛋白、V-set及免疫球蛋白区域包含蛋白4、Ifapsoriasin2、肌动蛋白胞质型1(Actin,cytoplasmic 1)、Suprabasin。
结论通过shotgun蛋白质组学技术以及GOA注释,我们获得人肺腺癌恶性胸腔积液详细的蛋白质组份信息,初步建立了人肺腺癌恶性胸腔蛋白质表达谱。123种高可信度蛋白质的绝大多数来源于血浆,只有7种蛋白质迄今为止尚未见报道在血浆中出现,包括膜联蛋白A2、JUP蛋白、转化生长因子βig-h3蛋白、V-set及免疫球蛋白区域包含蛋白4、Ifapsoriasin2、肌动蛋白胞质型1(Actin,cytoplasmic1)、Suprabasin,这些蛋白质存在于恶性胸腔积液而不易在血浆中被检测出来,因此可能有潜力成为诊断肺腺癌的分子标志物,其中膜联蛋白A2因其具有调节细胞的粘附和生长增殖能力而更具特殊意义。
Objective To acquire protein profiles of malignant pleural effusion from lung adenocarcinoma patients, provide information on the composition of protein contents in human malignant pleural effusion, in order to detect the potential candidates of useful biomarkers with diagnostic value in the pleural effusion of lung adenocarcinoma.
Method Pleural effusion samples were collected from five lung adenocarcinoma patients, and a composite sample was analyzed by using shotgun strategy (one-dimensional gel electrophoresis liquid chromatography-tandem mass spectrometry,1D-LC-MS/MS). Pleural effusion samples were separated by means of Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE ) and commassie blue staining .Proteomic analysis was performed by LC-MS/MS using an LTQ linear IT mass spectrometer (Thermo. San Jose, CA, USA),then the proteins were identified by using SEQUEST software and protein database search .And we classified all of the proteins identified in this study according to Gene Ontology Annotation (GOA) in aspects of molecule function, Subcellular location, and biological process as well.
Results Applying the shotgun strategy, we identified in total 230 unique proteins from human pleural effusion of lung adenocarcinoma patients. We classified these 230 proteins identified in this study according to the GOA, all of them were performed functional classification, biological process classification and Subcellular location classification. 123 proteins were identified with higher confidence levels (at least two unique peptide sequences matched),while 107 other proteins were identified at the minimal confidence level (only one unique peptide sequence matched). Most of these 123 proteins that were identified with higher confidence levels have been reported in plasma , however, among these 123 proteins,7 proteins , JUP protein, Suprabasin,AnnexinA2,Transforming growth factor-beta-induced protein ig-h3 precursor ,V-set and immunoglobulin domain-containing protein 4 precursor , Ifapsoriasin 2 ,and Actin, cytoplasmic1 have not been reported in plasma ,and they may represent proteins specifically present in pleural effusion.
Conclusion By using shotgun proteomic approaches and gene ontologyannotation (GOA), we have got fundamental information on the composition of protein contents in human malignant pleural effusion and profiled the protein express of human pleural effusion from lung adenocarcinoma patients. Most of these 123 proteins identified with higher confidence levels from human pleural effusion of lung adenocarcinoma patients are originated from plasma. However, 7 proteins, such as JUP protein, Suprabasin, Annexin A2,Transforming growth factor-beta-induced protein ig-h.3 precursor ,V-set and immunoglobulin domain-containing protein 4 precursor , Ifapsoriasin 2 and Actin,cytoplasmic1have not been reported in plasma. These proteins could represent potential candidates of useful biomarkers that could not be readily detected in plasma but in malignant pleural effusion. AnnexinA2 is of special interest since it may play a role in the regulation of intercellular adhesion and cell proliferation.
引文
[1]陈灏珠,主编.实用内科学[M].第12版.北京:人民卫生出版社,2005,1757.
[2]Valdes L,Alvarez D,Valle JM,et al.The etiology of pleural effusions in an area with high incidence of tuberculosis[J].Chest,1996,109:158-162.
[3]Marel M,Zrustova M,Stasny B,Light RW.The incidence ofpleural effusion in a well-defined region:epidemiologic study in central Bohemia[J].Chest,1993,104:1486-1489.
[4]Pakin DM,Pisani P,Feriary J,et al.Global cancer statistics[J].CA Cancer J Clin,1999,49:33-64.
[5]Granville CA,Dennis PA.An overview of lung cancer genomics and proteomics[J].Am J Respir Cell Mol Biol,2005,32(3):169-176.
[6]DeCamp MM,Mentzer SJ,Swanson SJ,et al.Malignant effusive disease of the pleura and pericardium.Chest,1997,112:291-295.
[7]Porcel JM,Vives M,Esquerda A,et al.Use of a panel of tumor markers(carcinoembryonic antigen,cancer antigen 125,caebohydrate antigen 15-3,and cytokeratin 19 fragments) in pleural fluid for the differential diagnosis of benign and malignant effusions.Chest,2004,126(6):1757-1763.
[8]Ando S,Suzuki M,Yamamoto N,et al.The prognostic value of both neuron-specific enzyme (NSE) and Cyfra21-1 in small cell lung cancer.Anticancer Res,2004,24(3b):1941-1946.
[9]Wilkins MR,Sanchez JC,Gooley AA,et al.Progress with proteome projects:why all proteins expressed by a genome should be identified and how to do it[J].Biotech& Genetic Engineering Rev,1996,13:41-50.
[10]Dove A.Proteomics:translating genomics into products? Nature Biotechnology,1999,17(3):233-236.
[11]Lee WC,Lee KH.Applications of affinity chromatography proteomics.Anal Biochem,2004,324(1):1-10.
[12]Wolters DA,Washburn MP,Yates JR.An automated multidimensional Protein identification technology for shotgun proteomics[J].Anal Chem,2001,73,5683-5690.
[13]Wu CC,MacCoss MJ,Howell KE,et al.A method for the comprehensive proteomic analysis of membrane Proteins[J].Nat Bio technol,2003,21,532-538.
[1]Aderson NL,Anderson NG.The human plasma proteome:history,character,and diagnostic prospects[J].Mol Cell Proteomics,2002,1(11),845-867.
[2]ManabeT,Muzuma H,Watanabe K.A nondenaturing protein map of human plasma proteins correlated with a denaturing polypeptide map combining techniques of micro two-dimensional gel electrophoresis[y].Electrophoresis,1999,20(4-5),830-835.
[3]Sloane AJ,DuffJL,WilsonNL,et al.High throughput peptide mass fmgerprinting and protein macro array analysis using chemical printing strategies[y].Proteomics Mol Cell,2002,1,490-499.
[4]Wu SL,Amato H,Biringer R,et al.Targeted proteomics of low-level proteins in human plasma by LC/MS:using human growth hormone as a model system[J].Proteome Res,2002,1,459-465.
[5]Choudhary G,Wu SL,Shieh P,et al.Multiple enzymatic digestion for enhanced sequence coverage of proteins in complex proteomic mixtures using capillary LC with ion trap MS/MS[J].Proteome Res,2003,2(1),59-67.
[6]Ulgiati D,Townend DC,Christiansen FT,et al.Complete sequence of the complement C4gene from the HLA-A1,B8,C4AQ0,C4B1,DR3 haplotype[J].Immunogenetics,1996,43:250-252.
[7]Anderson NL,Polanski M,Pieper R,et al.The human plasma proteome:a nonredundant list developed by combination of four separate sources[J].Mol Cell Proteomics,2004,3(4),311-326.
[8]Golaz O,Hughes GJ,Frutiger S,et al.Plasma and red blood cell protein maps:update 1993[J].Electrophoresis,1993,14(11),1223-1231.
[9]Hughes GJ,Frutiger S,Paquet N,et al.Plasma protein map:an update by microsequencing[J].Electrophoresis,1992,13(9-10),707-714.
[10]Pieper R,Gatlin CL,Makusky AJ,et al.The human serum proteome:display of nearly 3700chromatographically separated protein spots on two-dimensional electrophoresis gels and identification of 325 distinct proteins[J].Proteomics,2003,3,1345-1364.
[11]Ueno I,Sakai T,Yamaoka M,et al.Analysis of blood plasma proteins in patients with Alzheimer's disease by two-dimensional electrophoresis,sequence homology and immunodetection[J].Electrophoresis,2000,21(9),1832-1845.
[12]Tirumalai RS,Chan KC,Prieto DA et al.Characterization of the low molecular weight human serum proteome[J].Mol Cell Proteomics,2003,2(10),1096-1103.
[13]Fujii K,Nakano T,Kawamura T.et al.Multidimensional protein profiling technology and its application to human plasma proteome[y].Proteome Res,2004,3,712-718.
[14]Bunkenborg J,Pilch BJ,Podtelejnikow AV,et al.Screening for N-glycosylated proteins by liquid chromatography mass spectrometry[J].Proteomics,2004.4:454-465.
[15]Ramachandran P,Boontheung P,Xie Y,et al.Identification of N-linked glycoproteins in human saliva by glycoprotein capture and mass spectrometry[J].Proteome Res,2006,5:1493-1503.
[16] Zhou GX, Chao L, Chao J. Kallistatin: a novel human tissue kallikrein inhibitor. Purification, characterization, and reactive center sequence [J].Biol Chem, 1992,267: 25873-25880.
[17] Liu T, Qian WJ, Gritsenko MA, et al. Human plasma N-glycoproteome analysis by immunoaffinity subtraction, hydrazide chemistry, and mass spectrometry. Proteome Res, 2005, 4:2070-2080.
[18] Takiguchi M, Haraguchi Y, Mori M. Human liver-type arginase gene: structure of the gene and analysis of the promoter region [J] .Nucleic Acids Res, 1988,16:8789 -8802.
[19] Illy C, Thielens NM, Gagnon J, et al. Effect of lactoperoxidase-catalyzed iodination on the Ca (2+)-dependent interactions of human C1s. Location of the iodination sites. Biochemistry, 1991,30:7135-7141.
[20] Rehli M, Krause SW, Andressen R. Molecular characterization of the gene for human cartilage gp-39 (CHI3L1), a member of the chitinase protein family and marker for late stages of macrophage differentiation[J].Genomics, 1997,43:221-225.
[21] Esworthy RS, ChuFF, Paxton RJ, et al. Characterization and partial amino acid sequence of human plasma glutathione peroxidase. Arch Biochem Biophys, 1991,286:330-336.
[22] He P, He ZH, Dai J, et al. The human plasma proteome: Analysis of Chinese serum using shotgun strategy [J].Proteomics 2005, 5, 1-12.
[23] www. plasmaproteomedatabase.org
[24] The MGC Project Team.The status, quality, and expansion of the NIH full-length cDNA project: the Mammalian Gene Collection (MGC).Genome Res, 2004,14:2121-2127.
[25] Clark HF, Gurney AL, Abaya E, et al. The secreted protein discovery initiative (SPDI), a large-scale effort to identify novel human secreted and transmembrane proteins: a bioinformatics assessment [J].Genome Res, 2003, 13:2265-2270.
[26] Jindal HK, Chaney WG, Anderson CW, et al. The protein-tyrosine kinase substrate, calpactin I heavy chain (p36), is part of the primer recognition protein complex that interacts with DNA polymerase alpha [J].Biol Chem, 991,266:5169-5176.
[27] Zhang Y, Wolf- Yadlin A, Ross PL, et al. Time-resolved mass spectrometry of tyrosine phosphorylation sites in the epidermal growth factor receptor signaling network reveals dynamic modules [J]. Mol Cell Proteomics ,2005,4:1240-1250.
[28] Olsen JV,Blagoev B,Gnad F,et al. Global, in vivo, and site-specific phosphorylation dynamics in signaling networks[J].Cell,2006,127:635-648.
[29] Chi A, Valencia JC, Hu ZZ, et al. Proteomic and bioinformatic characterization of the biogenesis and function of melanosomes[J].Proteome Res,2006,5:3135-3144.
[30] Langnaese K, Colleaux L, Kloos DU,et al .Cloning of Z39Ig, a novel gene with immunoglobulin-like domains located on human chromosome X[J].Biochim Biophys Acta,2000,1492:522-525.
[31] Wu Z. The human intermediate filament-associated protein family. Submitted (NOV-2004) to the EMBL/GenBank/DDBJ databases.
[32] Procaccio V, Salaza G, Ono S, et al. A mutation of beta-actin that alters depolymerization dynamics is associated with autosomal dominant developmental malformations, deafness, and dystonia[J].Am J Hum Genet,2006,78:947-960.
[33] Stueven T, Hartmann E, Goerlich D. Exportin 6: a novel nuclear export receptor that is specific for profiling. actin complexes[J].EMBO,22:5928-5940(2003).
[34] Wilkins MR, Sanchez JC, Gooley AA, et al. Progress with proteome projects: why all proteins expressed by a genome should be identified and how to do it[J].Biotech& Genetic Engineering Rev,1996,13:41-50.
[35]Joshua N,Susan M,Kenneth J,et al.Toward a human blood serum proteome:analysis by multi dimensional separation coupled with mass spectrometry.Molecular and cellular proteomics,2002,1(12):947-955.
[36]Wolters DA,Washburn MP,Yates JR.An automated multidimensional Protein identification technology for shotgun proteomics[J].Anal Chem,2001,73,5683-5690.
[37]Wu CC,MacCoss MJ,Howell KE,et al.A method for the comprehensive proteomic analysis of membrane Proteins[J].Nat Biotechnol,2003,21,532-538.
[38]Washburn MP,Wolters D,Yates JR,et al.Large scale analysis of the yeast proteome by multi dimensional protein identification technology[J].Nature biotechnology,2001,19:242-247.
[39]Porcel JM,Vives M,Esquerda A,et al.Use of Panel of Markers in Pleural Fluid for Differential Diagnosis of Benign and Malignant Effusions[J].Chest,2004,1 26(6):1757-1763.
[40]Wu WG,Tang XM,Hu W,et al.Identification and validation of metastasis-associated proteins in head and neck cancer cell lines by two-dimensional electrophoresis and mass Spectrometry[J].Clin Exp Metastasis,2002,19:319-326.
[41]Danielsen EM,Van Deurs B,Hansen GH."Nonclassical" secretion of annexin A2 to the lumenal side of the enterocyte brush border membrane[J].Biochemistry,2003,42(49):14670-14676.
[42]Farber SA,De Rose RA,Olson ES,et al.The zebrafish annexin gene family[J].Genome Res,2003,13(6A):1082-1096.
[43]MacLeod TJ,Kwon M,Filipenko N R,Waisman D M.Phospholipid-associated annexin A2-S100A 10 heterotetramer and its subunits:characterization of the interaction with tissue plasminogen activator,plasminogen,and plasmin[J].Biol Chem,2003,278(28):25577-25584.
[44]Xiong X D,Xu L Y,Shen Z Y,et al.Identification of differentially expressed proteins between human esophageal immortalized and carcinomatous cell lines by two-dimensional electrophoresis and MALDI-TOF-mass spectrometry[J].World Gastroenterology,2002,8(5):777-781.
[45]李萃,肖志强,章晓鹏,等.人肺鳞癌组织的血清蛋白质组学的比较分析[J].中国生物化学与分子生物学报,2005,21(3),357-368.
[46]Skonier J,Neubauer M,Madisen L,et al.cDNA cloning and sequence analysis ofβig-h3,a novel gene induced in a human adenocarcinoma cell line after treatment with transforming growth factor-beta[J].DNA Cell Biol,1992,11(7):511-522.
[47]Skonier J,Bennett K,Rothwell V,et al.βig-h3:a transforming growth factor-βresponsive gene encoding a secreted protein that inhibits cell attachment in vitro and suppresses the growth of CHO cells in nude mice[J].DNA Cell Biol,1994,13(6):571-584.
[48]Billings PC,Herrick DJ,Kucich U,et al.Extracellular matrix and nuclear localization ofβig-h3 in human bladder smooth muscle and fibroblast cells[J].Cell Bio chem,2000,79(2):261-273.
[1]Nowak R.Entering the postgenome era[J].Science,1995,270.
[2]Krishna RG,Wold F.Post-translational modification of proteins[J].Adv Enzymol Relat Areas Mol Biol,1993,67:265-298.
[3]陈主初,梁宋平,主编.肿瘤蛋白质组学[M].长沙:湖南科学技术出版社,2002,168.
[4]查人俊,主编.现代肺癌诊断与治疗[M].北京:人民军医出版社,1993,245.
[5]Rodriguez-Panadero F,Borderas Naranjo F,Lopez Mejias J.Pleural metastatic tumours and effusions:frequency and pathogenic mechanisms in a post-mortem series[J].Eur Respir J,1989,2(4):366-369.
[6]Wilkins MR,Sanchez JC,Gooley AA,et al.Progress with proteome projects:why all proteins expressed by a genome should be identified and how to do it[J].Biotech& Genetic Engineering Rev,1996,13:41-50.
[7]Dove A.Proteomics:translating genomics into products? Nature Biotechnology,1999,17(3):233-236.
[8]李佰良.功能蛋白质组学[J].生命的化学.1998,18(6):1-3.
[9]成海平,钱小红.蛋白质组研究的技术体系及其进展[J].生物化学与生物物理进展,2000,27(6):584-588.
[10]O'Farrell PH,High resolution two—dimensional electrophoresis of proteins[J].The Journal of Biological Chemistry,1975,250(10):4007-4021.
[11]Righetti PG,Castagna A,Herbert B,et al.Prefraction Techniques in proteome analysis[J].Proteomics,2003,3(8):1397-1407.
[12]Bienvenut WV,Sanchez JC,KarmimeA,et al.Toward a clinical molecular scanner for proteome research:parallel protein chemical processing before and during western blot.Anal Chem,1999,71:4800-4807.
[13]Gygi SP,Rist B,Gerber SA,et al.Quantitative analysis of complex protein mixtures using isotope-coded affinity tags[J].Nature Biotechnology,1999,17(10):994-999.
[14]Unlu M,Margan ME,Minden JS,Difference gel elecrophoresis:a single gel method for detecting changes in protein extracts.Electrophoresis,1997,18:2071-2077.
[15]Tonge R,Shaw J,Middleton B,et al.Validation and development of fluorescence twodimensional differential gel electrophoresis is proteomics technology[J].Proteomics,2001,1(3):377-396.
[16]Chambers G,Lawrie L,Cash P,et al.Proteomics:A new approach to the study of disease[J].Journal of Pathology,2000,192(3):280-288.
[17]Rabilloud T.Silver staining of 2-D electrophoresis gels.Methods Mol Bio,1999,112:297-305.
[18]Matsui NM.Staining of preparative 2-D gels:Coomassie blue and imidazole - zinc negative staining.Methods Mol Bio,1999,112:307-311.
[19]Berggren K,Chernokalskaya E,Steinberg TH,et al.Backgroundfree,high-sensitivity staining of proteins in one-and two-dimensional sodium dodecyl sulfate polyacrylamide gels using a luminescent ruthenium complex.Electrophoresis,2000,21:2509.
[20]Lopez MF,Berggren K,Chernokalskaya E.A comparison of silver stain and SYPRO Ruby Protein Gel Stain with respect to protein detection in two-dimensional gels and identification by peptide mass profiling.Electrophoresis,2000,21(17):3673-3683.
[21]Celis JE,Ostergaard M,Jensen NA,et al.Human and mouse proteomic databases:novel resources in the protein universe[J].FEBS Lett,1998,430(1-2):64-72.
[22]Michael J,Simon J.Advances in mass spectrometry for proteome analysis[J].Current Opinion in Biotechnology,2000,11:384-390.
[23]Fenn JB,Mann M,Meng CK,et al.Electro spray ionization for mass spectrometry of large biomolecules.Science,1989,246:64-71.
[24]Karas M,Hillenkamp F.Laser Desorption ionization of proteins with molecular masses exceeding 10 000 Daltons.Anal Chem,1988,60:2299-2301.
[25]Mann M,Wilm M.Error to lerant identification of peptides in sequence databases by peptide sequence tags[J].Anal Chem,1994,66(24):4390-4399.
[26]Fung ET,Wright GL,Dalmasso EA.Proteomic strategies for biomarker identication:progress and challenges[J].Curr Opn Mol Ther,2002,2(4):643-650.
[27]von Eggeling F,Junken K,Feide W,et al.Mass spectrometry meets chip technology:a new proteomic tool in cancer research[J]? Electrophorisis,2001,22:2898-2902.
[28]Wolters DA,Washburn M P,Yates JR.An automated multidimensional Protein identification technology for shotgun proteomics[J].Anal.Chem,2001,73,5683-5690.
[29]Wu CC,MacCoss MJ,Howell KE,et al.A method for the comprehensive proteomic analysis of membrane Proteins[J].Nat B iotechnol,2003,21,532-538.
[30]Porcel JM,Vives M,Esquerda A,et al.Use of Panel of Markers in Pleural Fluid for Differential Diagnosis of Benign and Malignant Effusions[J].Chest,2004,126(6):1757-1763.
[31]Wulfkuhle JD,Liotta LA,Pericoin EF.Proteomic applications for the early detection of cancer[J].Nat Rev Cancer,2003,3(4):267-275.
[32]Albertini RJ.Biomarker responses in human populations:towards a worldwide map[J].Mutation Res,1999,428(1-2):217-226.
[33]陈灏珠,主编.实用内科学[M].第12版.北京:人民卫生出版社,2005,1757.
[34]Valdes L,Alvarez D,Valle JM,et al.The etiology of pleural effusions in an area with high incidence of tuberculosis[J].Chest,1996,109:158-162.
[35]Marel M,Zrustova M,Stasny B,Light RW.The incidence of pleural effusion in a well-defined region:epidemiologic study in central Bohemia[J].Chest,1993,104:1486-1489.
[36]DeCamp MM,Mentzer SJ,Swanson SJ,et al.Malignant effusive disease of the pleura and pericardium.Chest,1997,112:291-295.
[37]Pothur RS,Mukesh V,Yinming Z,et al.Proteomics for cancer biomarker discovery[J].Clinical Chemistry,2002,48(8):1160-1169.
[38]Lee WC,Lee KH.Applications of affinity chromatography in proteomics[J].Anal Biochem,2004,324(1):1-10.
[39]Tyan YC,Wu HY,Lai WW,et al.Proteomic profiling of human pleural effusion using two-dimensional nano liquid chromatography tandem mass spectrometry[J].Journal of Proteome Research,2005,4(4):1274-1286.
[40]Tyan YC,Wu HY,Su WC,et al.Proteomic analysis of human pleural effusion[J].Proteomics,2005,5(4):1062-1074.
[41]Hsieh WY,Chen MW,Ho HT,et al.Identification of differentially expressed proteins in human malignant pleural effusions[J].European Respiratory Journal,2006,28(6):1178-1185.
[42]Bard MP,Hegmans JP,Hemmes A,et al.Proteomic analysis of exosomes isolated from human malignant pleural effusions[J].American Journal of Respiratory Cell and Molecular Biology,2004,31(1):114-121.
[43]Davidson B,Espina V,Steinberg SM,et al.Proteomic analysis of malignant ovarian cancer effusions as a tool for biologic and prognostic profiling[J].Clinical Cancer Research,2006,12(31):791-799.
[44]吴广平,侯伟健,周海涛,等.双向凝胶电泳分析诊断肺癌性胸腔积液的临床意义[J].中华肿瘤防治杂志,2006,3(4):318-319.
[2]Valdes L,Alvarez D,Valle JM,et al.The etiology of pleural effusions in an area with high incidence of tuberculosis[J].Chest,1996,109:158-162.
[3]Marel M,Zrustova M,Stasny B,Light RW.The incidence ofpleural effusion in a well-defined region:epidemiologic study in central Bohemia[J].Chest,1993,104:1486-1489.
[4]Pakin DM,Pisani P,Feriary J,et al.Global cancer statistics[J].CA Cancer J Clin,1999,49:33-64.
[5]Granville CA,Dennis PA.An overview of lung cancer genomics and proteomics[J].Am J Respir Cell Mol Biol,2005,32(3):169-176.
[6]DeCamp MM,Mentzer SJ,Swanson SJ,et al.Malignant effusive disease of the pleura and pericardium.Chest,1997,112:291-295.
[7]Porcel JM,Vives M,Esquerda A,et al.Use of a panel of tumor markers(carcinoembryonic antigen,cancer antigen 125,caebohydrate antigen 15-3,and cytokeratin 19 fragments) in pleural fluid for the differential diagnosis of benign and malignant effusions.Chest,2004,126(6):1757-1763.
[8]Ando S,Suzuki M,Yamamoto N,et al.The prognostic value of both neuron-specific enzyme (NSE) and Cyfra21-1 in small cell lung cancer.Anticancer Res,2004,24(3b):1941-1946.
[9]Wilkins MR,Sanchez JC,Gooley AA,et al.Progress with proteome projects:why all proteins expressed by a genome should be identified and how to do it[J].Biotech& Genetic Engineering Rev,1996,13:41-50.
[10]Dove A.Proteomics:translating genomics into products? Nature Biotechnology,1999,17(3):233-236.
[11]Lee WC,Lee KH.Applications of affinity chromatography proteomics.Anal Biochem,2004,324(1):1-10.
[12]Wolters DA,Washburn MP,Yates JR.An automated multidimensional Protein identification technology for shotgun proteomics[J].Anal Chem,2001,73,5683-5690.
[13]Wu CC,MacCoss MJ,Howell KE,et al.A method for the comprehensive proteomic analysis of membrane Proteins[J].Nat Bio technol,2003,21,532-538.
[1]Aderson NL,Anderson NG.The human plasma proteome:history,character,and diagnostic prospects[J].Mol Cell Proteomics,2002,1(11),845-867.
[2]ManabeT,Muzuma H,Watanabe K.A nondenaturing protein map of human plasma proteins correlated with a denaturing polypeptide map combining techniques of micro two-dimensional gel electrophoresis[y].Electrophoresis,1999,20(4-5),830-835.
[3]Sloane AJ,DuffJL,WilsonNL,et al.High throughput peptide mass fmgerprinting and protein macro array analysis using chemical printing strategies[y].Proteomics Mol Cell,2002,1,490-499.
[4]Wu SL,Amato H,Biringer R,et al.Targeted proteomics of low-level proteins in human plasma by LC/MS:using human growth hormone as a model system[J].Proteome Res,2002,1,459-465.
[5]Choudhary G,Wu SL,Shieh P,et al.Multiple enzymatic digestion for enhanced sequence coverage of proteins in complex proteomic mixtures using capillary LC with ion trap MS/MS[J].Proteome Res,2003,2(1),59-67.
[6]Ulgiati D,Townend DC,Christiansen FT,et al.Complete sequence of the complement C4gene from the HLA-A1,B8,C4AQ0,C4B1,DR3 haplotype[J].Immunogenetics,1996,43:250-252.
[7]Anderson NL,Polanski M,Pieper R,et al.The human plasma proteome:a nonredundant list developed by combination of four separate sources[J].Mol Cell Proteomics,2004,3(4),311-326.
[8]Golaz O,Hughes GJ,Frutiger S,et al.Plasma and red blood cell protein maps:update 1993[J].Electrophoresis,1993,14(11),1223-1231.
[9]Hughes GJ,Frutiger S,Paquet N,et al.Plasma protein map:an update by microsequencing[J].Electrophoresis,1992,13(9-10),707-714.
[10]Pieper R,Gatlin CL,Makusky AJ,et al.The human serum proteome:display of nearly 3700chromatographically separated protein spots on two-dimensional electrophoresis gels and identification of 325 distinct proteins[J].Proteomics,2003,3,1345-1364.
[11]Ueno I,Sakai T,Yamaoka M,et al.Analysis of blood plasma proteins in patients with Alzheimer's disease by two-dimensional electrophoresis,sequence homology and immunodetection[J].Electrophoresis,2000,21(9),1832-1845.
[12]Tirumalai RS,Chan KC,Prieto DA et al.Characterization of the low molecular weight human serum proteome[J].Mol Cell Proteomics,2003,2(10),1096-1103.
[13]Fujii K,Nakano T,Kawamura T.et al.Multidimensional protein profiling technology and its application to human plasma proteome[y].Proteome Res,2004,3,712-718.
[14]Bunkenborg J,Pilch BJ,Podtelejnikow AV,et al.Screening for N-glycosylated proteins by liquid chromatography mass spectrometry[J].Proteomics,2004.4:454-465.
[15]Ramachandran P,Boontheung P,Xie Y,et al.Identification of N-linked glycoproteins in human saliva by glycoprotein capture and mass spectrometry[J].Proteome Res,2006,5:1493-1503.
[16] Zhou GX, Chao L, Chao J. Kallistatin: a novel human tissue kallikrein inhibitor. Purification, characterization, and reactive center sequence [J].Biol Chem, 1992,267: 25873-25880.
[17] Liu T, Qian WJ, Gritsenko MA, et al. Human plasma N-glycoproteome analysis by immunoaffinity subtraction, hydrazide chemistry, and mass spectrometry. Proteome Res, 2005, 4:2070-2080.
[18] Takiguchi M, Haraguchi Y, Mori M. Human liver-type arginase gene: structure of the gene and analysis of the promoter region [J] .Nucleic Acids Res, 1988,16:8789 -8802.
[19] Illy C, Thielens NM, Gagnon J, et al. Effect of lactoperoxidase-catalyzed iodination on the Ca (2+)-dependent interactions of human C1s. Location of the iodination sites. Biochemistry, 1991,30:7135-7141.
[20] Rehli M, Krause SW, Andressen R. Molecular characterization of the gene for human cartilage gp-39 (CHI3L1), a member of the chitinase protein family and marker for late stages of macrophage differentiation[J].Genomics, 1997,43:221-225.
[21] Esworthy RS, ChuFF, Paxton RJ, et al. Characterization and partial amino acid sequence of human plasma glutathione peroxidase. Arch Biochem Biophys, 1991,286:330-336.
[22] He P, He ZH, Dai J, et al. The human plasma proteome: Analysis of Chinese serum using shotgun strategy [J].Proteomics 2005, 5, 1-12.
[23] www. plasmaproteomedatabase.org
[24] The MGC Project Team.The status, quality, and expansion of the NIH full-length cDNA project: the Mammalian Gene Collection (MGC).Genome Res, 2004,14:2121-2127.
[25] Clark HF, Gurney AL, Abaya E, et al. The secreted protein discovery initiative (SPDI), a large-scale effort to identify novel human secreted and transmembrane proteins: a bioinformatics assessment [J].Genome Res, 2003, 13:2265-2270.
[26] Jindal HK, Chaney WG, Anderson CW, et al. The protein-tyrosine kinase substrate, calpactin I heavy chain (p36), is part of the primer recognition protein complex that interacts with DNA polymerase alpha [J].Biol Chem, 991,266:5169-5176.
[27] Zhang Y, Wolf- Yadlin A, Ross PL, et al. Time-resolved mass spectrometry of tyrosine phosphorylation sites in the epidermal growth factor receptor signaling network reveals dynamic modules [J]. Mol Cell Proteomics ,2005,4:1240-1250.
[28] Olsen JV,Blagoev B,Gnad F,et al. Global, in vivo, and site-specific phosphorylation dynamics in signaling networks[J].Cell,2006,127:635-648.
[29] Chi A, Valencia JC, Hu ZZ, et al. Proteomic and bioinformatic characterization of the biogenesis and function of melanosomes[J].Proteome Res,2006,5:3135-3144.
[30] Langnaese K, Colleaux L, Kloos DU,et al .Cloning of Z39Ig, a novel gene with immunoglobulin-like domains located on human chromosome X[J].Biochim Biophys Acta,2000,1492:522-525.
[31] Wu Z. The human intermediate filament-associated protein family. Submitted (NOV-2004) to the EMBL/GenBank/DDBJ databases.
[32] Procaccio V, Salaza G, Ono S, et al. A mutation of beta-actin that alters depolymerization dynamics is associated with autosomal dominant developmental malformations, deafness, and dystonia[J].Am J Hum Genet,2006,78:947-960.
[33] Stueven T, Hartmann E, Goerlich D. Exportin 6: a novel nuclear export receptor that is specific for profiling. actin complexes[J].EMBO,22:5928-5940(2003).
[34] Wilkins MR, Sanchez JC, Gooley AA, et al. Progress with proteome projects: why all proteins expressed by a genome should be identified and how to do it[J].Biotech& Genetic Engineering Rev,1996,13:41-50.
[35]Joshua N,Susan M,Kenneth J,et al.Toward a human blood serum proteome:analysis by multi dimensional separation coupled with mass spectrometry.Molecular and cellular proteomics,2002,1(12):947-955.
[36]Wolters DA,Washburn MP,Yates JR.An automated multidimensional Protein identification technology for shotgun proteomics[J].Anal Chem,2001,73,5683-5690.
[37]Wu CC,MacCoss MJ,Howell KE,et al.A method for the comprehensive proteomic analysis of membrane Proteins[J].Nat Biotechnol,2003,21,532-538.
[38]Washburn MP,Wolters D,Yates JR,et al.Large scale analysis of the yeast proteome by multi dimensional protein identification technology[J].Nature biotechnology,2001,19:242-247.
[39]Porcel JM,Vives M,Esquerda A,et al.Use of Panel of Markers in Pleural Fluid for Differential Diagnosis of Benign and Malignant Effusions[J].Chest,2004,1 26(6):1757-1763.
[40]Wu WG,Tang XM,Hu W,et al.Identification and validation of metastasis-associated proteins in head and neck cancer cell lines by two-dimensional electrophoresis and mass Spectrometry[J].Clin Exp Metastasis,2002,19:319-326.
[41]Danielsen EM,Van Deurs B,Hansen GH."Nonclassical" secretion of annexin A2 to the lumenal side of the enterocyte brush border membrane[J].Biochemistry,2003,42(49):14670-14676.
[42]Farber SA,De Rose RA,Olson ES,et al.The zebrafish annexin gene family[J].Genome Res,2003,13(6A):1082-1096.
[43]MacLeod TJ,Kwon M,Filipenko N R,Waisman D M.Phospholipid-associated annexin A2-S100A 10 heterotetramer and its subunits:characterization of the interaction with tissue plasminogen activator,plasminogen,and plasmin[J].Biol Chem,2003,278(28):25577-25584.
[44]Xiong X D,Xu L Y,Shen Z Y,et al.Identification of differentially expressed proteins between human esophageal immortalized and carcinomatous cell lines by two-dimensional electrophoresis and MALDI-TOF-mass spectrometry[J].World Gastroenterology,2002,8(5):777-781.
[45]李萃,肖志强,章晓鹏,等.人肺鳞癌组织的血清蛋白质组学的比较分析[J].中国生物化学与分子生物学报,2005,21(3),357-368.
[46]Skonier J,Neubauer M,Madisen L,et al.cDNA cloning and sequence analysis ofβig-h3,a novel gene induced in a human adenocarcinoma cell line after treatment with transforming growth factor-beta[J].DNA Cell Biol,1992,11(7):511-522.
[47]Skonier J,Bennett K,Rothwell V,et al.βig-h3:a transforming growth factor-βresponsive gene encoding a secreted protein that inhibits cell attachment in vitro and suppresses the growth of CHO cells in nude mice[J].DNA Cell Biol,1994,13(6):571-584.
[48]Billings PC,Herrick DJ,Kucich U,et al.Extracellular matrix and nuclear localization ofβig-h3 in human bladder smooth muscle and fibroblast cells[J].Cell Bio chem,2000,79(2):261-273.
[1]Nowak R.Entering the postgenome era[J].Science,1995,270.
[2]Krishna RG,Wold F.Post-translational modification of proteins[J].Adv Enzymol Relat Areas Mol Biol,1993,67:265-298.
[3]陈主初,梁宋平,主编.肿瘤蛋白质组学[M].长沙:湖南科学技术出版社,2002,168.
[4]查人俊,主编.现代肺癌诊断与治疗[M].北京:人民军医出版社,1993,245.
[5]Rodriguez-Panadero F,Borderas Naranjo F,Lopez Mejias J.Pleural metastatic tumours and effusions:frequency and pathogenic mechanisms in a post-mortem series[J].Eur Respir J,1989,2(4):366-369.
[6]Wilkins MR,Sanchez JC,Gooley AA,et al.Progress with proteome projects:why all proteins expressed by a genome should be identified and how to do it[J].Biotech& Genetic Engineering Rev,1996,13:41-50.
[7]Dove A.Proteomics:translating genomics into products? Nature Biotechnology,1999,17(3):233-236.
[8]李佰良.功能蛋白质组学[J].生命的化学.1998,18(6):1-3.
[9]成海平,钱小红.蛋白质组研究的技术体系及其进展[J].生物化学与生物物理进展,2000,27(6):584-588.
[10]O'Farrell PH,High resolution two—dimensional electrophoresis of proteins[J].The Journal of Biological Chemistry,1975,250(10):4007-4021.
[11]Righetti PG,Castagna A,Herbert B,et al.Prefraction Techniques in proteome analysis[J].Proteomics,2003,3(8):1397-1407.
[12]Bienvenut WV,Sanchez JC,KarmimeA,et al.Toward a clinical molecular scanner for proteome research:parallel protein chemical processing before and during western blot.Anal Chem,1999,71:4800-4807.
[13]Gygi SP,Rist B,Gerber SA,et al.Quantitative analysis of complex protein mixtures using isotope-coded affinity tags[J].Nature Biotechnology,1999,17(10):994-999.
[14]Unlu M,Margan ME,Minden JS,Difference gel elecrophoresis:a single gel method for detecting changes in protein extracts.Electrophoresis,1997,18:2071-2077.
[15]Tonge R,Shaw J,Middleton B,et al.Validation and development of fluorescence twodimensional differential gel electrophoresis is proteomics technology[J].Proteomics,2001,1(3):377-396.
[16]Chambers G,Lawrie L,Cash P,et al.Proteomics:A new approach to the study of disease[J].Journal of Pathology,2000,192(3):280-288.
[17]Rabilloud T.Silver staining of 2-D electrophoresis gels.Methods Mol Bio,1999,112:297-305.
[18]Matsui NM.Staining of preparative 2-D gels:Coomassie blue and imidazole - zinc negative staining.Methods Mol Bio,1999,112:307-311.
[19]Berggren K,Chernokalskaya E,Steinberg TH,et al.Backgroundfree,high-sensitivity staining of proteins in one-and two-dimensional sodium dodecyl sulfate polyacrylamide gels using a luminescent ruthenium complex.Electrophoresis,2000,21:2509.
[20]Lopez MF,Berggren K,Chernokalskaya E.A comparison of silver stain and SYPRO Ruby Protein Gel Stain with respect to protein detection in two-dimensional gels and identification by peptide mass profiling.Electrophoresis,2000,21(17):3673-3683.
[21]Celis JE,Ostergaard M,Jensen NA,et al.Human and mouse proteomic databases:novel resources in the protein universe[J].FEBS Lett,1998,430(1-2):64-72.
[22]Michael J,Simon J.Advances in mass spectrometry for proteome analysis[J].Current Opinion in Biotechnology,2000,11:384-390.
[23]Fenn JB,Mann M,Meng CK,et al.Electro spray ionization for mass spectrometry of large biomolecules.Science,1989,246:64-71.
[24]Karas M,Hillenkamp F.Laser Desorption ionization of proteins with molecular masses exceeding 10 000 Daltons.Anal Chem,1988,60:2299-2301.
[25]Mann M,Wilm M.Error to lerant identification of peptides in sequence databases by peptide sequence tags[J].Anal Chem,1994,66(24):4390-4399.
[26]Fung ET,Wright GL,Dalmasso EA.Proteomic strategies for biomarker identication:progress and challenges[J].Curr Opn Mol Ther,2002,2(4):643-650.
[27]von Eggeling F,Junken K,Feide W,et al.Mass spectrometry meets chip technology:a new proteomic tool in cancer research[J]? Electrophorisis,2001,22:2898-2902.
[28]Wolters DA,Washburn M P,Yates JR.An automated multidimensional Protein identification technology for shotgun proteomics[J].Anal.Chem,2001,73,5683-5690.
[29]Wu CC,MacCoss MJ,Howell KE,et al.A method for the comprehensive proteomic analysis of membrane Proteins[J].Nat B iotechnol,2003,21,532-538.
[30]Porcel JM,Vives M,Esquerda A,et al.Use of Panel of Markers in Pleural Fluid for Differential Diagnosis of Benign and Malignant Effusions[J].Chest,2004,126(6):1757-1763.
[31]Wulfkuhle JD,Liotta LA,Pericoin EF.Proteomic applications for the early detection of cancer[J].Nat Rev Cancer,2003,3(4):267-275.
[32]Albertini RJ.Biomarker responses in human populations:towards a worldwide map[J].Mutation Res,1999,428(1-2):217-226.
[33]陈灏珠,主编.实用内科学[M].第12版.北京:人民卫生出版社,2005,1757.
[34]Valdes L,Alvarez D,Valle JM,et al.The etiology of pleural effusions in an area with high incidence of tuberculosis[J].Chest,1996,109:158-162.
[35]Marel M,Zrustova M,Stasny B,Light RW.The incidence of pleural effusion in a well-defined region:epidemiologic study in central Bohemia[J].Chest,1993,104:1486-1489.
[36]DeCamp MM,Mentzer SJ,Swanson SJ,et al.Malignant effusive disease of the pleura and pericardium.Chest,1997,112:291-295.
[37]Pothur RS,Mukesh V,Yinming Z,et al.Proteomics for cancer biomarker discovery[J].Clinical Chemistry,2002,48(8):1160-1169.
[38]Lee WC,Lee KH.Applications of affinity chromatography in proteomics[J].Anal Biochem,2004,324(1):1-10.
[39]Tyan YC,Wu HY,Lai WW,et al.Proteomic profiling of human pleural effusion using two-dimensional nano liquid chromatography tandem mass spectrometry[J].Journal of Proteome Research,2005,4(4):1274-1286.
[40]Tyan YC,Wu HY,Su WC,et al.Proteomic analysis of human pleural effusion[J].Proteomics,2005,5(4):1062-1074.
[41]Hsieh WY,Chen MW,Ho HT,et al.Identification of differentially expressed proteins in human malignant pleural effusions[J].European Respiratory Journal,2006,28(6):1178-1185.
[42]Bard MP,Hegmans JP,Hemmes A,et al.Proteomic analysis of exosomes isolated from human malignant pleural effusions[J].American Journal of Respiratory Cell and Molecular Biology,2004,31(1):114-121.
[43]Davidson B,Espina V,Steinberg SM,et al.Proteomic analysis of malignant ovarian cancer effusions as a tool for biologic and prognostic profiling[J].Clinical Cancer Research,2006,12(31):791-799.
[44]吴广平,侯伟健,周海涛,等.双向凝胶电泳分析诊断肺癌性胸腔积液的临床意义[J].中华肿瘤防治杂志,2006,3(4):318-319.