本文選題：EF-Tu + 多蛋白復(fù)合物��； 參考：《廈門大學(xué)》2007年碩士論文

【摘要】： 細(xì)菌延伸因子Tu (elongation factor Tu, EF-Tu)是一種高豐度蛋白,在蛋白質(zhì)生物合成的延伸過程中起到了關(guān)鍵性的作用。近年來,EF-Tu卻是作為一種新發(fā)現(xiàn)的病原相關(guān)分子模式(Pathogen-associated molecular patterns, PAMPs)分子而受到高度重視。作為一個(gè)具有高度保守性的菌細(xì)胞內(nèi)的蛋白質(zhì),它是如何從細(xì)胞內(nèi)跨膜運(yùn)動(dòng)到細(xì)胞外,并且被宿主細(xì)胞特異性地識(shí)別進(jìn)而引發(fā)宿主的免疫反應(yīng)的。同時(shí),在此過程中又有哪些膜蛋白起了重要的作用。這無疑具有著相當(dāng)重要的生物學(xué)意義。 試驗(yàn)首先采用Ni-NTA柱純化和生物質(zhì)譜技術(shù),以模式大腸桿菌為試驗(yàn)材料,發(fā)現(xiàn)了內(nèi)膜蛋白MdtE、外膜蛋白OmpF和具有4個(gè)跨膜片段的外膜蛋白OmpA與漿蛋白EF-Tu之間存在著蛋白質(zhì)相互作用,形成了1個(gè)多蛋白復(fù)合體,被命名為EF-Tu多蛋白復(fù)合體。同時(shí)采用Western blotting、Far-Western blotting和免疫共沉淀等技術(shù),充分證實(shí)了上述結(jié)論。并提出了該多蛋白質(zhì)復(fù)合體的分子結(jié)構(gòu)模型。在此基礎(chǔ)上,研究這4種蛋白在胞漿中的結(jié)合情況。結(jié)果發(fā)現(xiàn),他們以EF-Tu·OmpA,OmpF·OmpA和MdtE·OmpA 3種兩兩結(jié)合的形式存在。 進(jìn)一步探討該復(fù)合體在細(xì)胞分泌性蛋白中的狀態(tài),發(fā)現(xiàn)該多蛋白質(zhì)復(fù)合體缺少了MdtE,而以EF-Tu、OmpA和OmpF蛋白復(fù)合體形式分泌至胞外。最后,對(duì)EF-Tu多蛋白復(fù)合體與人單核/巨噬細(xì)胞結(jié)合情況進(jìn)行了研究,發(fā)現(xiàn)EF-Tu、OmpA和OmpF仍然以一個(gè)多蛋白復(fù)合體的形式和細(xì)胞發(fā)生了相互作用,其中EF-Tu和OmpA為直接結(jié)合分子。這些結(jié)果提示OmpA和OmpF在EF-Tu被宿主細(xì)胞識(shí)別和引起免疫反應(yīng)的過程中發(fā)揮了重要作用。 上述研究首次發(fā)現(xiàn)了EF-Tu多蛋白復(fù)合體,揭示了該復(fù)合體從胞漿至胞外的形成和運(yùn)輸特征,闡明了其與動(dòng)物細(xì)胞作用形式。這些研究結(jié)果對(duì)深入了解EF-Tu病原相關(guān)分子模式的生物學(xué)功能具有重要作用。
[Abstract]:The bacterial elongation factor Tub (EF-TuS) is a high abundance protein, which plays a key role in the extension of protein biosynthesis. Recently, EF-Tu has attracted much attention as a novel pathogen-associated molecular pattern (PAMPs) molecule. As a highly conserved intracellular protein, it moves from the intracellular membrane to the outside of the cell and is specifically recognized by the host cell to trigger the host's immune response. At the same time, which membrane proteins play an important role in this process. There is no doubt that this is of great biological significance. Firstly, Ni-NTA column purification and biomass spectrometry were used in the experiment, and the model Escherichia coli was used as the experimental material. The protein interaction between membrane protein MdtE, outer membrane protein OmpF and outer membrane protein OmpA with four transmembrane fragments and plasma protein EF-Tu was found, resulting in the formation of a polyprotein complex named EF-Tu polyprotein complex. The results were confirmed by Western blotting Far-Western blotting and immunoprecipitation. The molecular structure model of the polyprotein complex was proposed. On this basis, the binding of these four proteins in the cytoplasm was studied. The results showed that they existed in the form of EF-Tu OmpF OmpA and MdtE OmpA. Further study of the state of the complex in cell secretory protein showed that the polyprotein complex lacked MdtE, but secreted to the extracellular in the form of EF-Tu OmpA and OmpF protein complex. Finally, the binding of EF-Tu polyprotein complex to human mononuclear / macrophage was studied. It was found that EF-Tu and OmpF still interacted with cells in the form of a polyprotein complex, in which EF-Tu and OmpA were direct binding molecules. These results suggest that OmpA and OmpF play an important role in the recognition and immune response of EF-Tu by host cells. The interaction with animal cells was elucidated. These results play an important role in understanding the biological functions of EF-Tu related molecular models.
【學(xué)位授予單位】：廈門大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2007
【分類號(hào)】：R378

【引證文獻(xiàn)】

相關(guān)博士學(xué)位論文 前2條

1 王方昆;馬耳他型布氏桿菌弱毒疫苗株M5-90致弱分子機(jī)制的研究[D];中國(guó)農(nóng)業(yè)科學(xué)院;2011年

2 梁廷明;中華絨螯蟹螺原體重要功能基因的篩選和研究及螺原體與WSSV的多重PCR檢測(cè)技術(shù)[D];南京師范大學(xué);2011年

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本文編號(hào)：2009267