本文選題：β2糖蛋白I + 乙型肝炎病毒��； 參考：《吉林大學(xué)》2009年碩士論文

【摘要】： 乙型肝炎病毒(HBV)是一種嗜肝脫氧核糖核酸病毒,目前全球約有4億人群感染HBV。多年來(lái)學(xué)者對(duì)HBV感染肝細(xì)胞的早期過(guò)程進(jìn)行研究,提出了多種觀點(diǎn)并對(duì)可能的靶細(xì)胞受體進(jìn)行鑒定,但仍無(wú)定論。β2糖蛋白I(β2-GPI)又稱為載脂蛋白H,是血漿中一種含量較豐富的糖蛋白。近年研究認(rèn)為,β2-GPI是抗磷脂抗體綜合征(APS)的主要抗原之一。APS是與抗磷脂抗體(aPL)密切相關(guān),是以血栓形成、習(xí)慣性流產(chǎn)和血小板減少等癥狀為特點(diǎn)的一組綜合征。β2-GPI與aPL結(jié)合后能識(shí)別帶負(fù)電荷的磷脂復(fù)合物,并使β2-GPI的表面結(jié)構(gòu)發(fā)生變化,使機(jī)體易形成血栓。隨著研究的拓展,Mehdi發(fā)現(xiàn)重組乙型肝炎表面抗原(rHBsAg)可與正常人肝細(xì)胞膜通過(guò)β2-GPI在膜外結(jié)合,表明β2-GPI可能參與HBV感染肝細(xì)胞。本研究組深入研究后提出,β2-GPI可能作為中介分子,與HBsAg形成復(fù)合物,再與肝細(xì)胞膜表面的特異性蛋白結(jié)合,從而介導(dǎo)HBV入肝過(guò)程。研究組已從肝癌細(xì)胞株SMMC-7721表面經(jīng)鑒定出結(jié)合蛋白為膜聯(lián)蛋白Ⅱ。在本研究中,我們將著重探討β2-GPI與HBsAg特異性結(jié)合的親和力及二者相互作用的機(jī)制,為上述觀點(diǎn)提供更有力的實(shí)驗(yàn)依據(jù),為今后的研究奠定基礎(chǔ)。本研究利用大腸桿菌M15(pQE30-hβ2-GPI)高效表達(dá)目的蛋白,通過(guò)親和層析純化帶有融合6個(gè)組氨酸標(biāo)簽的β2-GPI包涵體,并進(jìn)行復(fù)性。原核系統(tǒng)表達(dá)的目的蛋白具有無(wú)糖基化修飾的特點(diǎn),故這種rβ2-GPI可用于研究分子構(gòu)象對(duì)蛋白結(jié)合作用的影響。我們首次利用放射性免疫法(radioimmunoassay,RIA)測(cè)定人β2GPI與rHBsAg結(jié)合的親和常數(shù)(Ka),以此推斷血漿中β2GPI與HBsAg的親和力,并通過(guò)親和常數(shù)比較與原核系統(tǒng)表達(dá)的rβ2-GPI免疫活性的差異。梯度濃度的兩組來(lái)源不同的β2GPI分別與125I-rHBsAg結(jié)合,利用標(biāo)準(zhǔn)曲線Adrion法分別測(cè)得兩組蛋白的Ka。采用嵌套實(shí)驗(yàn)設(shè)計(jì)進(jìn)行數(shù)據(jù)分析,血漿中提取的β2-GPI組與原核系統(tǒng)表達(dá)的rβ2-GPI組的Ka無(wú)明顯統(tǒng)計(jì)學(xué)差異(P0.05)。最終求得血漿人β2-GPI組:Ka1=(2.795±1.846)×10~8 L/ mol。結(jié)果表明血漿β2-GPI與HBsAg結(jié)合的親和力較強(qiáng),且血漿中提取的β2-GPI與原核系統(tǒng)表達(dá)的rβ2-GPI與rHBsAg的結(jié)合力相似。我們得出結(jié)論,血漿中β2-GPI與HBsAg親和力強(qiáng),可能在血漿中二者較易形成復(fù)合物,且二者的結(jié)合與β2GPI的糖基化結(jié)構(gòu)無(wú)關(guān),即糖基化與否不影響β2-GPI的免疫活性。因此,β2-GPI無(wú)論發(fā)生何種構(gòu)象改變,均不影響其與HBsAg的結(jié)合。這種結(jié)合可能參與介導(dǎo)HBV嗜肝過(guò)程,甚至參與肝細(xì)胞癌的發(fā)生發(fā)展。
[Abstract]:Hepatitis B virus (HBV) is a hepatophilic deoxyribonucleic acid virus. At present, about 400 million people worldwide are infected with HBV. Over the years, researchers have studied the early process of HBV infection in hepatocytes, put forward a variety of viewpoints and identified possible target cell receptors. 尾 2 glycoprotein I (尾 2 GPI), also known as apolipoprotein H, is a rich glycoprotein in plasma. In recent years, it has been considered that 尾 2-GPI is one of the major antigens of anti-phospholipid antibody syndrome (APSs). APS is closely related to anti-phospholipid antibody (2-GPI). A group of syndromes characterized by habitual abortion and thrombocytopenia. 尾 2-GPI combined with aPL can recognize the phospholipid complex with negative charge and change the surface structure of 尾 2-GPI and make the body easily form thrombosis. With the development of the study, Mehdi found that recombinant hepatitis B surface antigen (HBs) could bind to normal human hepatocytes through 尾 2-GPI, suggesting that 尾 2-GPI may be involved in the infection of hepatocytes with HBV. This study suggests that 尾 2-GPI may act as a mediator, form a complex with HBsAg, and then bind to specific proteins on the surface of the hepatocyte membrane, thus mediating the process of HBV entering the liver. The binding protein was identified as integrin 鈪，

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