本文選題：人胎肝 + 細胞質(zhì)膜��； 參考：《中國人民解放軍軍事醫(yī)學(xué)科學(xué)院》2005年博士論文

【摘要】：細胞質(zhì)膜構(gòu)成細胞對外界環(huán)境的屏障和細胞內(nèi)外環(huán)境交流的界面,鑲嵌或連接于其中的蛋白質(zhì)對維持細胞正常的生理活動具有十分重要的作用,這些蛋白質(zhì)參與細胞—細胞或細胞—細胞外基質(zhì)的識別、粘附、連接和通訊,信號的接受和跨膜傳導(dǎo),細胞內(nèi)外物質(zhì)的轉(zhuǎn)運等,影響著細胞的生長、發(fā)育、分化、病變等。細胞質(zhì)膜蛋白占到已知藥物靶標的70%,因此,細胞質(zhì)膜已經(jīng)成為廣泛的藥物設(shè)計的靶標,細胞質(zhì)膜蛋白質(zhì)組得到學(xué)術(shù)界的高度重視。 16-24周孕齡人胎肝處于關(guān)鍵的造血調(diào)控和發(fā)育分化時期,分析人胎肝細胞質(zhì)膜蛋白質(zhì)組有助于對人胎肝的這些生理活動的認識。我們用差速離心和密度梯度離心提取純化了人胎肝細胞質(zhì)膜,細胞質(zhì)膜標志酶—5’單核苷酸酶特異酶活測定顯示其富集度達到16倍。用SDS-PAGE-RPLC-ESI-MS/MS和“shotgun”(蛋白三重裂解—SCX—RPLC-ESI—Q-TOF-MS)兩條技術(shù)路線,我們從人胎肝細胞膜和堿處理細胞膜中鑒定了總共532個非冗余蛋白質(zhì),另有82個蛋白group。這是目前已知最大的人類組織細胞質(zhì)膜表達譜。在人胎肝細胞質(zhì)膜蛋白質(zhì)表達譜中,已知明確定位于細胞質(zhì)膜上的蛋白質(zhì)126個,占鑒定蛋白總數(shù)的24.5%�？缒さ鞍�162個,占30.5%,未知蛋白41個。126個細胞質(zhì)膜蛋白基本覆蓋了細胞質(zhì)膜上所有的功能模塊,其中轉(zhuǎn)運體、膜骨架、信號轉(zhuǎn)導(dǎo)、受體、細胞連接、細胞粘附和代謝酶類蛋白所占比例最高,分別為18、15、14、11、10、9和6%。這與人胎肝細胞質(zhì)膜的結(jié)構(gòu)和生理功能密切相關(guān)。另外,在人胎肝細胞質(zhì)膜上鑒定了一些已報道在細胞質(zhì)膜上有定位但常見于其它亞細胞結(jié)構(gòu)的蛋白,如細胞質(zhì)蛋白HSP27和HSP70;內(nèi)質(zhì)網(wǎng)蛋白GRP和PDI;線粒體蛋白VDAC-1、HSP60和ATP合成酶;細胞核蛋白Lasp-1,hRNP和組蛋白。這些蛋白質(zhì)可能參與新的細胞質(zhì)膜上的生理活動。 鑒于在提取人胎肝細胞質(zhì)膜時不可避免地存在其它內(nèi)膜系統(tǒng)的污染,對于鑒定的非已知細胞質(zhì)膜蛋白,我們不能確定是發(fā)生真正的定位變化而存在于細胞質(zhì)膜還是其它亞細胞結(jié)構(gòu)的污染。正常人紅細胞只由胞漿和細胞膜構(gòu)成,因而紅細胞膜不會有其它亞細胞結(jié)構(gòu)的污染。所以我們選用正常人紅細胞膜作為模式細胞膜進一步對其蛋白質(zhì)組進行分析。通過與研究人胎肝細胞質(zhì)膜相同的
[Abstract]:The cytoplasmic membrane forms the barrier of cell to the outside environment and the interface of the communication between the inside and outside of the cell. The protein embedded or connected to it plays a very important role in maintaining the normal physiological activities of the cell. These proteins are involved in the recognition, adhesion, connection and communication of cell-cell or cell-extracellular matrix (ECM), signal reception and transmembrane transduction, and the transport of substances inside and outside the cell, which affect cell growth, development, differentiation, pathological changes and so on. Cytoplasmic membrane proteins account for 70 percent of the known drug targets. Therefore, cytoplasmic membrane has become a widely used target for drug design. Human fetal liver at 16-24 weeks of gestational age is in a critical stage of hematopoietic regulation and differentiation. Analyzing the proteome of plasma membrane of human fetal liver cell is helpful to the understanding of these physiological activities of human fetal liver. The plasma membrane of human fetal hepatocytes was extracted and purified by differential centrifugation and density gradient centrifugation. Using SDS-PAGE-RPLC-ESI-MS/MS and "shotgun" (triple cleavage of protein, SCX-RPLC-ESI-Q-TOF-MS), a total of 532 non-redundant proteins were identified from human fetal liver cell membrane and alkali-treated cell membrane, and 82 protein groups were identified. This is the largest known expression profile of the plasma membrane of human tissue cells. In the plasma membrane protein expression profile of human fetal liver cells, 126 proteins were known to be located on the cytoplasmic membrane, accounting for 24.5% of the total identified proteins. Transmembrane proteins 162 (30.5%), unknown proteins 41. 126 cytoplasmic membrane proteins basically cover all functional modules on the cytoplasmic membrane, including transporters, membrane cytoskeletons, signal transduction, receptors, cell junctions. The proportion of cell adhesion and metabolizing enzyme proteins was the highest, which was 1815, 141110109 and 6g, respectively. This is closely related to the structure and physiological function of the plasma membrane of human fetal hepatocytes. In addition, some proteins, such as cytoplasmic proteins HSP27 and HSP70, endoplasmic reticulum proteins GRP and PDI, mitochondrial proteins VDAC-1HSP60 and ATP synthase, were identified on the plasma membrane of human fetal hepatocytes. Nuclear protein Lasp-1 hRNP and histone. These proteins may be involved in physiological activities on the new cytoplasmic membrane. In view of the inevitable contamination of other intimal systems in the extraction of the plasma membrane of human fetal hepatocytes, the identification of non-known cytoplasmic membrane proteins, We are not sure whether a real localization change occurs in the cytoplasmic membrane or other subcellular structure contamination. Normal human erythrocytes consist only of cytoplasm and cell membrane, so there is no other subcellular structure contamination in erythrocyte membrane. So we selected the normal human erythrocyte membrane as the model cell membrane to further analyze its proteome. The same as the study of the plasma membrane of human fetal hepatocytes
【學(xué)位授予單位】：中國人民解放軍軍事醫(yī)學(xué)科學(xué)院
【學(xué)位級別】：博士
【學(xué)位授予年份】：2005
【分類號】：R341

【引證文獻】

相關(guān)博士學(xué)位論文 前1條

1 汪磊;肝癌導(dǎo)向肽結(jié)合蛋白的篩選及其功能研究[D];華東師范大學(xué);2007年

相關(guān)碩士學(xué)位論文 前1條

1 王振玲;牛精子質(zhì)膜提取及二維電泳分離精子全蛋白的技術(shù)研究[D];內(nèi)蒙古農(nóng)業(yè)大學(xué);2006年

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本文編號：1928434