流行性感冒病毒有著極高的罹病率，死亡率及造成重大的經濟負擔，科學家們都致力於發展能快速診斷及對抗流感病毒的材料。麥芽糖結合蛋白能與amylose結合，常被做為載體來增加重組蛋白在大腸桿菌中的溶解性。甘露糖結合凝集素屬於凝集素家族，是由肝臟所製造的類膠原質的血清蛋白，C端有碳水化合物辨識區域(CRD)，負責與碳水化合物作用，文獻報導它能與流感病毒的血球凝集素與神經胺酸酶結合。在我的研究中，利用這兩種蛋白來測試阻斷流感病毒感染的能力。先利用RT-PCR從Huh7細胞株得到MBL2的CRD cDNA後，將其轉接到pRSET載體。然後在大腸桿菌中表現重組蛋白MBP和MBL-CRD，再分別經由amylose resin及polyhistidine-tagged親和性層析做純化。首先藉由ELISA來確認MBP及MBL-CRD與流感病毒結合的能力，我們也發現加入MBP和MBL-CRD能夠阻斷流感病毒感染的機會而提高MDCK細胞的存活率，而加入mannose能廢止MBL-CRD的保護效果。然而將MBP與金奈米接合後，它的生物性功能便顯著性的增強，但MBL-CRD接上金奈米卻沒有這樣的效果，甚至修飾上hetero-PEG也沒有改善。之後我們會利用動物實驗進一步證實MBP和MBL-CRD的保護效果。最後，我們期望Au-MBP和Au-CRD能成為發展對流感病毒的診斷及治療上的潛力材料。

Influenza epidemics result in significant morbidity, mortality, and economic burden. Development of rapid diagnosis and novel anti-viral strategies is important to combat influenza. Maltose binding protein (MBP), known to bind amylose, is used to increase the solubility of recombinant proteins expressed in E. coli. Mannose-binding lectin (MBL), a liver-derived collagen-like serum protein that has been reported to bind to hemagglutinin (HA) and neuraminidase (NA) of influenza A virus (IAV), belongs to collectin family and has a carbohydrate recognition domain (CRD) at C-terminus, which is responsible for interactions with carbohydrate patterns on non-self-surfaces. In this study, we used two proteins to test their abilities of blocking influenza virus infection. We obtained the CRD cDNA of MBL2 from Huh7 cells using RT-PCR and cloned it into the pRSET prokaryotic expression vector. Recombinant MBP and MBL-CRD proteins were produced in E. coli and purified by amylose resin and the polyhistidine-tagged affinity chromatography, respectively. First, we confirmed the binding activity of MBP and MBL-CRD to IAV by ELISA. We also found that treatment of MBP and MBL-CRD could increase the viability of MDCK cells infected with IAV. The protection effect of MBL-CRD was abolished by mannose. Strikingly, the biological function of MBP was amplified after conjugating with gold nanoparticle (AuNP). However, MBL-CRD conjugated with AuNP did not amplify its function, even when it was modified with polyethylene glycol (PEG). We will prove the protection effect of MBP and MBL-CRD in animal challenge models. In conclusion, our results suggest that Au-MBP and Au-CRD may be further explored for developing new diagnostic and therapeutic agents for influenza virus.

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