Microsoft Word doc

Size: px

Start display at page:

Download "Microsoft Word doc"

서헉 송
5 years ago
Views:

1 KISEP Rhinology Korean J Otolaryngol 2006;49:41-6 프로테오믹스 (Proteomics) 를이용한반전성유두종의특이발현단백질분석 이재용 1 이상학 2 Proteomic Analysis of the Specific Protein Expression in Inverted Papilloma Jae Yong Lee, MD 1 and Sang Hag Lee, MD 2 1 Department of Otorhinolaryngology-Head and Neck Surgery, Soonchunhyang University College of Medicine, Bucheon and 2 Department of Otorhinolaryngology-Head and Neck Surgery, Korea University College of Medicine, Seoul, Korea ABSTRACT Background and Objectives:Inverted papilloma is a benign tumor of nasal cavity and paranasal sinuses with a propensity for local invasiveness, recurrence, and malignant transformation. Proteomics is a powerful tool for protein analysis, providing valuable information on biochemical processes involved in diseases, monitoring of cellular processes, and characterizing the protein expression levels. We tried to find the proteins that are associated with pathophysiology of the inverted papilloma and mechanisms of the disease by proteomic approach. Materials and Method:Normal nasal mucosa and inverted papilloma tissue was obtained during augmentation rhinoplasty and endoscopic surgery, respectively. Total protein was isolated and separated into numerous spots by two-dimensional electrophoresis. Twenty four protein spots that were only detected in inverted papilloma were selected and subsequently analyzed with matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS). Results:About 700 protein spots were detected. Selected spots were analyzed, and various proteins were identified. These include T-cell receptor beta chains, Ca 2+ binding proteins, caltractin, calneuron, ras-related proteins, a rab-2b oncogene family, chloride intracellular channel proteins, tumor protein D53, and tumor necrosis factor precursors. Conclusion:We identified the proteins expressed in the inverted papilloma with proteomic approach. These proteins may help us in understanding the mechanisms of pathogenesis of inverted papilloma, and may be used as possible tumor markers. (Korean J Otolaryngol 2006;49:41-6) KEY WORDS:Proteomics Inverted papilloma. - 41

2 반전성유두종의특이발현단백질분석 Korean J Otolaryngol 2006;49:41-6

3 이재용외 Table 1. Identified proteins that were only expressed in the inverted papilloma tissue. Protein name, sequence coverage rates, number of matched peptides, accession code, isoelectric point and molecular weight are listed Spot No. Protein name Sequence coverage rates No. of matched peptides Accession code pim.wt 01 T-cell receptor beta chain Apolipoprotein C-II precursor Ca 2+ binding protein Myosin regulatory light chain P Caltractin Q Calneuron Q9BXU Ras-related protein RAP-2b P RAP-2b, member of ras oncogene family Sodium channel beta-3 subunit precursor Q9NY Chloride intracellular channel protein Interleukin-1 alpha precursor P G protein signaling regulator Tumor protein D Q Ras-related protein RAP-1b P Immunoglobulin lambda light chain Proteasome activator complex subunit Q MHC class I antigen Tumor necrosis factor precursor P Interleukin-6 precursor P Ras-related protein Rab-33B Q9H Immunoglobulin heavy chain variable region Alcohol sulfotransferase Q Dual specificity mitogen-activated protein kinase Serologically defined colon cancer antigen pi : isoelectric point, MWt : molecular weight

31,000 고 찰 21,500 14,400 프로테옴(proteome)이란 단어는 어원적으로 단백질체라 6,500 고 풀이될 수 있으나 실제적으로는 게놈(gemone)의 상대 어로서 프로테오믹스는 이러한 protein과 genomics의 합 성어라 할 수 있다.

4 반전성 유두종의 특이 발현 단백질 분석 MWt (Da) 데, 일반적으로 5개 이상의 일치하는 펩타이드와 15% 이 200,000 상의 염기서열차지율을 가진 단백질을 선택하게 된다. 본 연 97,400 구결과 동정된 단백질은 이에 합당한 조건을 가지고 있었다. 66,200 조직을 효소처리하여 분석하면 아직까지 밝혀지지 않거나 명 45,000 tical protein 또는 unmanned protein product라 칭하게 명되지 않은 단백질들도 동정될 수 있는데 이는 hypothe되며 본 연구에서는 이러한 단백질들은 동정되지 않았다. 31,000 고 찰 21,500 14,400 프로테옴(proteome)이란 단어는 어원적으로 단백질체라 6,500 고 풀이될 수 있으나 실제적으로는 게놈(gemone)의 상대 어로서 프로테오믹스는 이러한 protein과 genomics의 합 성어라 할 수 있다.10-12) 이 단어는 1995년 이탈리아 Siena pi 4 7 에서 개최된 이차원적 전기영동학회(2-dimensional elec- Fig. 1. Two-dimensional electrophoresis gel of normal human nasal mucosa. The proteins from the inferior turbinate mucosa were extracted and separated on ph 4to 7 IPG strips, followed by 12% polyacrylamide gel electrophoresis. The gel was stained with silver nitrate. Above 4.5 kda region, protein spots were clouded making a protein band, and below this many scattered protein spots can be detected. trophoresis meeting)에서 Marc Wilkins에 의해 처음 사 용된 이후 보편화되었다. 게놈, 즉 유전체는 각 종(species) 마다 동일하며 정적인 성질을 띄는 반면, 단백질은 어떠한 개체에 주어진 환경에 따라 능동적으로 발현되며 해당 유 전자 없이도 특정 단백질이 발현되거나 유전자의 존재에도 MWt (Da) 200,000 불구하고 단백질이 발현되지 않을 수 있다. 더욱이 유전자 전이후변형이나 단백질간의 상호작용에 의해 전혀 새로운 단 97,400 백질의 형성도 가능하기 때문에 유전체에 대한 연구에서 단 66,200 백질체로의 연구로 관심이 전환된 이유가 여기에 있다고 할 수 있다.13) 프로테오믹스는 크게 이차원적 전기영동과 질량 45,000 분석기를 통한 단백질동정으로 구성되며, 이 중 이차원적 전 기영동은 조직이나 체액내의 단백질을 분리하고 그 발현정 도를 알 수 있는 방법으로서 프로테오믹스의 근간을 이루 , , , ,500 MS 외에 좀더 정확한 단백질의 동정이 가능한 기계들의 발 전에 의존하여 프로테오믹스는 최근 급성장하고 있다.17)18) 저자들은 프로테오믹스를 이용하여 정상비점막과 반전 성 유두종에서 발현되는 단백질을 알아보고 이러한 단백질 을 비교검토함으로써 가능한 종양표지자의 발견과 함께 반 21 전성 유두종 발생에 관여하는 병태생리학적 기전을 이해하 pi 4 는 기술이다.5-10) 또한 염색기술의 발달과 MALDI-TOF 7 고자 하였다. MALDI-TOF MS를 이용하여 반전성 유두 Fig. 2. Two-dimensional electrophoresis gel of inverted papilloma. Gross pattern of protein spot expression is similar to that of the normal nasal mucosa. Using melanie III software, we detected protein spots that were only expressed in the inverted papilloma. Marked number shows the selected protein spots. 종에서만 특이적으로 발현되는 단백질반점들을 분석하여 동 는 경향을 보이고 있다. 동정된 단백질에 대한 신뢰도는 일 chain, MHC class I antigen, immunoglobulin lambda light 치하는 펩타이드의 수(number of matched peptides)와 염 chain, immunoglobulin heavy chain variable region 등 기서열차지율(sequence coverage rates)에 의해 결정되는 면역조절에 관여하는 단백질, ras-related protein RAP- 44 정된 단백질들은 Table 1과 같다. 동정된 단백질들에 대하 여 살펴보면 caltractin, calneuron, Ca2+ binding protein 등 칼슘조절에 관여하는 단백질들과 T-cell receptor beta Korean J Otolaryngol 2006;49:41-6

5 이재용외 REFERENCES 1) Han JK, Smith TL, Loehrl T, Toohill RJ, Smith MM. An evolution in the management of sinonasal inverting papilloma. Laryngoscope 2001; 111: ) Kaufman MR, Brandwein MS, Lawson W. Sinonasal papillomas: Clinicopathologic review of 40 patients with inverted and oncocytic schneiderian papillomas. Laryngoscope 2002;112: ) Thorp MA, Oyarzabal-Amigo MF, du Plessis JH, Sellars SL. Inverted papilloma: A review of 53 cases. Laryngoscope 2001;111: ) Hwang CS, Yang HS, Hong MK. Detection of human papillomavirus (HPV) in sinonasal inverted papillomas using polymerase chain reaction (PCR). Am J Rhinol 1998;12: ) Beranova-Giorgianni S, Giorgianni F, Desiderio DM. Analysis of the proteome in the human pituitary. Proteomics 2002;2: ) Langen H, Berndt P, Roder D, Cairns N, Lubec G, Fountoulakis M. Two-dimensional map of human brain proteins. Electrophoresis 1999; 20: ) Beranova-Giorgianni S, Pabst MJ, Russell TM, Giorgianni F, Goldowitz D, Desiderio DM. Preliminary analysis of the mouse cerebellum proteome. Brain Res Mol Brain Res 2002;98: ) Jenkins RE, Pennington SR. Arrays for protein expression profiling: Toward a viable alternative to two-dimensional gel electrophoresis? Proteomics 2001;1: ) Zuo X, Speicher DW. Comprehensive analysis of complex proteomes using microscale solution isoelectric focusing prior to narrow ph range two-dimensional electrophoresis. Proteomics 2002;2: ) Zuo X, Speicher DW. A method for global analysis of complex proteomes using sample prefractionation by solution isoelectric focusing prior to two-dimensional electrophoresis. Anal Biochem 2000;284: ) Tan C, Li J, Wang J, Xiang Q, Zhang X, Dong L, et al. Proteomic analysis of differential protein expression in human nasopharyngeal carcinoma cells induced by NAG7 transfection. Proteomics 2002;2: ) Park KS, Kim H, Kim NG, Cho SY, Choi KH, Seong JK, et al. Proteomic analysis and molecular characterization of tissue ferritin light chain in hepatocellular carcinoma. Hepatology 2002;35: ) Ahram M, Best CJ, Flaig MJ, Gillespie JW, Leiva IM, Chuaqui RF, et al. Proteomic analysis of human prostate cancer. Mol Carcinog 2002; 33:

6 반전성유두종의특이발현단백질분석 14) Tsuji T, Shimohama S, Kamiya S, Sazuka T, Ohara O. Analysis of brain proteins in Alzheimer s disease using high-resolution two-dimensional gel electrophoresis. J Neurol Sci 1999;166: ) Lim SO, Park SJ, Kim W, Park SG, Kim HJ, Kim YI, et al. Proteome analysis of hepatocellular carcinoma. Biochem Biophys Res Commun 2002;291: ) Lauber WM, Carroll JA, Dufield DR, Kiesel JR, Radabaugh MR, Malone JP. Mass spectrometry compatibility of two-dimensional gel protein stains. Electrophoresis 2001;22: ) Knepper MA. Proteomics and kidney. J Am Soc Nephrol 2002;13: ) Beranova-Giorgianni S, Desiderio DM. Mass spectrometry of the human pituitary proteome: Identification of selected proteins. Rapid Commun Mass Spectrom 2000;14: ) Herbert B. Advances in protein solubilization for two-dimensional electrophoresis. Electrophoresis 1999;20: ) Gevaert K, Demoi H, Martens L, Hoorelbeke B, Puype M, Goethals M, et al. Protein identification based on matrix assisted laser desorption/ionization-post source decay-mass spectrometry. Electrophoresis 2001;22: Korean J Otolaryngol 2006;49:41-6

Can032.hwp

Can032.hwp Chromosomal Alterations in Hepatocellular Carcinoma Cell Lines Detected by Comparative Genomic Hybridization Sang Jin Park 1, Mahn Joon Ha, Ph.D. 1, Hugh Chul Kim, M.D. 2 and Hyon Ju Kim, M.D. 1 1 Laboratory