淀粉样蛋白致病机理及其调控的体外研究
|
摘要
淀粉样蛋白的错误折叠和异常聚集是2型糖尿病、阿兹海默病、海绵状脑病等淀粉样变性病的重要致病原因之一。淀粉样蛋白的细胞毒性主要来源于淀粉样蛋白的聚集、氧化压力的产生以及与细胞膜的相互作用；细胞毒性的产生过程会受到翻译后修饰、金属离子、共存蛋白以及二维细胞膜表面等多种因素的调控。在本论文中，我们重点研究了磷酸化和泛素化等翻译后修饰、金属铜离子、二维负电表面等因素对淀粉样蛋白细胞毒性产生的调控机理，并同时着重对蛋白定点泛素化合成的新方法进行了探索。本研究对理解淀粉样变性病病理机理具有重要意义，且为相应的治疗手段和药物的开发提供了理论基础。
我们首先利用高效原子力显微镜和石英微天平技术，对胰岛淀粉样多肽(hIAPP)和胰岛素(Insulin)在五氧化二钽表面以不同比例共聚集的过程和形貌特征进行了研究，结果表明不同比率的hIAPP/Insulin在负电表面形成不同的聚集纳米结构，表现为成熟纤维-寡聚体纤维-寡聚体-前体原纤维-环状纤维等形式。石英晶体微天平表征结果显示蛋白共混后对表面的吸附作用更强。研究结果对进一步研究2型糖尿病的病理机理有重要的参考价值。
我们利用多肽固相合成方法以及BC法、Ellman法和3-CCA法等荧光检测技术对Tau核心片段R3以及Ser324位磷酸化的片段pR3与铜离子作用产生氧化压力情况进行了研究。结果表明R3和pR3都具有还原铜的能力，R3和pR3在铜离子的介导下可以产生羟基自由基，而自身被氧化成由二硫键相连的二聚体，磷酸化会明显加速由R3与铜离子共同介导的活性氧物种产生的速率，这种加速效应会受到pH环境的影响，研究结果有助于我们更深入理解异常磷酸化在Tau病理学中的调控作用。
此外，我们通过表达蛋白连接技术和抑制基因引发突变技术，成功获得了泛素蛋白硫酯，并通过生物表达方法将含叠氮基的非天然氨基酸成功引入到泛素蛋白单体的63位之中；我们利用泛素蛋白硫酯的高反应活性在泛素单体C端成功引入了巯基基团和双键基团；利用光催化的巯基－烯基加成反应将含双键的泛素蛋白Ub-AA与含巯基的小分子MESNA进行了偶联；我们利用DNTB试剂作为辅助基团，成功将泛素蛋白单体Ub-SH与淀粉样相关蛋白Prion聚集核心片段prp175-195通过二硫键进行偶联。研究结果为进一步建立具有多位点翻译后修饰蛋白质的合成方法以及生物功能的研究奠定了基础。
Misfolding and abnormal aggregation of amyloid protein is an important cause forthe amyloidosis such as type2diabetes, Alzheimer’s disease and spongiformencephalopathy. The amyloid cytotoxicity mainly derived from the aggregation ofamyloid protein,the generation of oxidative stress, as well as the interaction with the cellmembrane. The process of cell toxicity generating will also be related withpost-translationally modified, the metal ions, the coexistence protein interactions as wellas a two-dimensional surface. In this paper, we focus on the effect of thephosphorylation and ubiquitination, metal copper ions, two-dimensional negativelycharged surface on the amyloid cytotoxicity.Moreover,we explored new methodology todesign and get the ubiquitinated protein.this will be helpful to understand the pathogenicmechanisms of the amyloidosis and develop new amyloidosis drugs.
We Use high resolution liquid AFM and QCM-D to investigate thenanostructures by co-assembling hIAPP/insulin on Ta2O5surfaces, we have shownthat the structure and morphology of co-assembled aggregates on negativelycharged surfaces depend on the ratio of hIAPP/insulin. By tuning thehIAPP/insulin ratio, the nanostructure morphology changes from fibrils to oligomers, toannular. These findings provide new insights to understand insulin’s affact onthe structure of hIAPP aggregates on biomembranes.
Furthermore, we investigated the generation of hydroxyl radicals when R3peptide was co-incubated with Cu(II). We also compared the redox activity ofR3with its phosphorylated form (pR3) at the site of Serine324. Phosphorylationat Ser324was found in paired helical filament in AD brains. Our resultsdemonstrated that both R3and pR3is able to reduce Cu(II). And the speed ofhydroxyl radical generation by R3and Cu(II) is significantly accelerated byphosphorylation. The effect of phosphorylation depends on pH condition.Thiseffect of phosphorylation on ROS generation by tau and copper implies a newperspective of its neurotoxicity.
In addition,we use the expressed protein ligation and suppressor mutagenesistechnology to successfully get the ubiquitin thioester,and we also get the ubiquitin monomer containing a non-natural amino acid in the63site. We take advantage of thehigh reactivity of ubiquitin thioester to introduce the mercapto group and double bondgroup to the C terminal of the ubiquitin monomer. the protein Ub-AA coupled with theMESNA through the thiol-ene radical addition reaction under photocatalyticcondition.We use the DTNB reagent as a leaving group to get the Ub-SH monomercoupling with the Prion fragment PrP175-195via a disulfide bond.
我们首先利用高效原子力显微镜和石英微天平技术，对胰岛淀粉样多肽(hIAPP)和胰岛素(Insulin)在五氧化二钽表面以不同比例共聚集的过程和形貌特征进行了研究，结果表明不同比率的hIAPP/Insulin在负电表面形成不同的聚集纳米结构，表现为成熟纤维-寡聚体纤维-寡聚体-前体原纤维-环状纤维等形式。石英晶体微天平表征结果显示蛋白共混后对表面的吸附作用更强。研究结果对进一步研究2型糖尿病的病理机理有重要的参考价值。
我们利用多肽固相合成方法以及BC法、Ellman法和3-CCA法等荧光检测技术对Tau核心片段R3以及Ser324位磷酸化的片段pR3与铜离子作用产生氧化压力情况进行了研究。结果表明R3和pR3都具有还原铜的能力，R3和pR3在铜离子的介导下可以产生羟基自由基，而自身被氧化成由二硫键相连的二聚体，磷酸化会明显加速由R3与铜离子共同介导的活性氧物种产生的速率，这种加速效应会受到pH环境的影响，研究结果有助于我们更深入理解异常磷酸化在Tau病理学中的调控作用。
此外，我们通过表达蛋白连接技术和抑制基因引发突变技术，成功获得了泛素蛋白硫酯，并通过生物表达方法将含叠氮基的非天然氨基酸成功引入到泛素蛋白单体的63位之中；我们利用泛素蛋白硫酯的高反应活性在泛素单体C端成功引入了巯基基团和双键基团；利用光催化的巯基－烯基加成反应将含双键的泛素蛋白Ub-AA与含巯基的小分子MESNA进行了偶联；我们利用DNTB试剂作为辅助基团，成功将泛素蛋白单体Ub-SH与淀粉样相关蛋白Prion聚集核心片段prp175-195通过二硫键进行偶联。研究结果为进一步建立具有多位点翻译后修饰蛋白质的合成方法以及生物功能的研究奠定了基础。
Misfolding and abnormal aggregation of amyloid protein is an important cause forthe amyloidosis such as type2diabetes, Alzheimer’s disease and spongiformencephalopathy. The amyloid cytotoxicity mainly derived from the aggregation ofamyloid protein,the generation of oxidative stress, as well as the interaction with the cellmembrane. The process of cell toxicity generating will also be related withpost-translationally modified, the metal ions, the coexistence protein interactions as wellas a two-dimensional surface. In this paper, we focus on the effect of thephosphorylation and ubiquitination, metal copper ions, two-dimensional negativelycharged surface on the amyloid cytotoxicity.Moreover,we explored new methodology todesign and get the ubiquitinated protein.this will be helpful to understand the pathogenicmechanisms of the amyloidosis and develop new amyloidosis drugs.
We Use high resolution liquid AFM and QCM-D to investigate thenanostructures by co-assembling hIAPP/insulin on Ta2O5surfaces, we have shownthat the structure and morphology of co-assembled aggregates on negativelycharged surfaces depend on the ratio of hIAPP/insulin. By tuning thehIAPP/insulin ratio, the nanostructure morphology changes from fibrils to oligomers, toannular. These findings provide new insights to understand insulin’s affact onthe structure of hIAPP aggregates on biomembranes.
Furthermore, we investigated the generation of hydroxyl radicals when R3peptide was co-incubated with Cu(II). We also compared the redox activity ofR3with its phosphorylated form (pR3) at the site of Serine324. Phosphorylationat Ser324was found in paired helical filament in AD brains. Our resultsdemonstrated that both R3and pR3is able to reduce Cu(II). And the speed ofhydroxyl radical generation by R3and Cu(II) is significantly accelerated byphosphorylation. The effect of phosphorylation depends on pH condition.Thiseffect of phosphorylation on ROS generation by tau and copper implies a newperspective of its neurotoxicity.
In addition,we use the expressed protein ligation and suppressor mutagenesistechnology to successfully get the ubiquitin thioester,and we also get the ubiquitin monomer containing a non-natural amino acid in the63site. We take advantage of thehigh reactivity of ubiquitin thioester to introduce the mercapto group and double bondgroup to the C terminal of the ubiquitin monomer. the protein Ub-AA coupled with theMESNA through the thiol-ene radical addition reaction under photocatalyticcondition.We use the DTNB reagent as a leaving group to get the Ub-SH monomercoupling with the Prion fragment PrP175-195via a disulfide bond.
引文
[1] Lazarov O, Mattson M P, Peterson D A, et al. When neurogenesis encounters aging anddisease. Trends Neurosci.2010,33(12):569-579.
[2] Opie E L. The relation oe diabetes mellitus to lesions of the pancreas. hyaline degeneration ofthe islands oe langerhans. J Exp Med.1901,5(5):527-540.
[3] Soto C. Unfolding the role of protein misfolding in neurodegenerative diseases. Nat RevNeurosci.2003,4(1):49-60.
[4] Pardes H, Manton K G, Lander E S, et al. Effects of medical research on health care andeconomy. Science.1999,283(5398):36-37.
[5] Zimmet P. The burden of type2diabetes: are we doing enough? Diabetes Metab.2003,29(4Pt2):6S-9S.
[6] Zhang S, Liu H, Yu H, et al. Fas-associated death receptor signaling evoked by human amylinin islet beta-cells. Diabetes.2008,57(2):348-356.
[7] Hoppener J W, Lips C J. Role of islet amyloid in type2diabetes mellitus. Int J Biochem CellBiol.2006,38(5-6):726-736.
[8] Butler A E, Janson J, Bonner-Weir S, et al. Beta-cell deficit and increased beta-cell apoptosisin humans with type2diabetes. Diabetes.2003,52(1):102-110.
[9] Ward B, Walker K, Exley C. Copper(II) inhibits the formation of amylin amyloid in vitro. JInorg Biochem.2008,102(2):371-375.
[10] Sloane P D, Zimmerman S, Suchindran C, et al. The public health impact of Alzheimer'sdisease,2000-2050: potential implication of treatment advances. Annu Rev Public Health.2002,23:213-231.
[11] Mandelkow E. Alzheimer's disease. The tangled tale of tau. Nature.1999,402(6762):588-589.
[12] Gong C X, Liu F, Grundke-Iqbal I, et al. Post-translational modifications of tau protein inAlzheimer's disease.[J]. J Neural Transm.2005,112(6):813-838.
[13] Buee L, Bussiere T, Buee-Scherrer V, et al. Tau protein isoforms, phosphorylation and role inneurodegenerative disorders. Brain Res Brain Res Rev.2000,33(1):95-130.
[14] Dobson C M. The structural basis of protein folding and its links with human disease. PhilosTrans R Soc Lond B Biol Sci.2001,356(1406):133-145.
[15] Cohen F E, Pan K M, Huang Z, et al. Structural clues to prion replication.Science.1994,264(5158):530-531.
[16]陈媚莎.淀粉样蛋白聚集的机理研究和抑制剂的设计、合成与评价[博士学位论文].北京:清华大学化学系.2012.
[17] Salmona M, Malesani P, De Gioia L, et al. Molecμlar determinants of the physicochemicalproperties of a critical prion protein region comprising residues106-126. Biochem J.1999,342(Pt1):207-214.
[18] De Gioia L, Selvaggini C, Ghibaudi E, et al. Conformational polymorphism of theamyloidogenic and neurotoxic peptide homologous to residues106-126of the prion protein. JBiol Chem.1994,269(11):7859-7862.
[19] Jobling M F, Stewart L R, White A R, et al. The hydrophobic core sequence modμlates theneurotoxic and secondary structure properties of the prion peptide106-126. J Neurochem.1999,73(4):1557-1565.
[20] Dupiereux I, Zorzi W, Lins L, et al. Interaction of the106-126prion peptide with lipidmembranes and potential implication for neurotoxicity. Biochem Biophys Res Commun.2005,331(4):894-901.
[21] Kayed R, Head E, Thompson J L, et al. Common structure of soluble amyloid oligomersimplies common mechanism of pathogenesis. Science.2003,300(5618):486-489.
[22] Zraika S, Hμll R L, Udayasankar J, et al. Oxidative stress is induced by islet amyloidformation and time-dependently mediates amyloid-induced beta cell apoptosis. Diabetologia.2009,52(4):626-635.
[23] Janson J, Ashley R H, Harrison D, et al. The mechanism of islet amyloid polypeptide toxicityis membrane disruption by intermediate-sized toxic amyloid particles. Diabetes.1999,48(3):491-498.
[24] Mirzabekov T A, Lin M C, Kagan B L. Pore formation by the cytotoxic islet amyloid peptideamylin. J Biol Chem.1996,271(4):1988-1992.
[25] Brundin P, Melki R, Kopito R. Prion-like transmission of protein aggregates inneurodegenerative diseases. Nat Rev Mol Cell Biol.2010,11(4):301-307.
[26] Khemtemourian L, Killian J A, Hoppener J W, et al. Recent insights in islet amyloidpolypeptide-induced membrane disruption and its role in beta-cell death in type2diabetesmellitus. Exp Diabetes Res.2008,2008:421287.
[27] Kudva Y C, Mueske C, Butler P C, et al. A novel assay in vitro of human islet amyloidpolypeptide amyloidogenesis and effects of insμlin secretory vesicle peptides on amyloidformation. Biochem J.1998,331(Pt3):809-813.
[28] Janciauskiene S, Eriksson S, Carlemalm E, et al. B cell granμle peptides affect human isletamyloid polypeptide (IAPP) fibril formation in vitro. Biochem Biophys Res Commun.1997,236(3):580-585.
[29] Cui W, Ma J W, Lei P, et al. Insμlin is a kinetic but not a thermodynamic inhibitor of amylinaggregation. FEBS J.2009,276(12):3365-3371.
[30] Zraika S, Hμll R L, Udayasankar J, et al. Oxidative stress is induced by islet amyloidformation and time-dependently mediates amyloid-induced beta cell apoptosis. Diabetologia.2009,52(4):626-635.
[31] Masad A, Hayes L, Tabner B J, et al. Copper-mediated formation of hydrogen peroxide fromthe amylin peptide: a novel mechanism for degeneration of islet cells in type-2diabetesmellitus? FEBS Lett.2007,581(18):3489-3493.
[32] Butterfield S M, Lashuel H A. Amyloidogenic protein-membrane interactions: mechanisticinsight from model systems. Angew Chem Int Ed Engl.2010,49(33):5628-5654.
[33] Brender J R, Durr U H, Heyl D, et al. Membrane fragmentation by an amyloidogenic fragmentof human Islet Amyloid Polypeptide detected by solid-state NMR spectroscopy of membranenanotubes. Biochim Biophys Acta.2007,1768(9):2026-2029.
[34] Knight J D, Miranker A D. Phospholipid catalysis of diabetic amyloid assembly. J Mol Biol.2004,341(5):1175-1187.
[35] Ambroggio E E, Kim D H, Separovic F, et al. Surface behavior and lipid interaction ofAlzheimer beta-amyloid peptide1-42: a membrane-disrupting peptide. Biophys J.2005,88(4):2706-2713.
[36] Kowalewski T, Holtzman D M. In situ atomic force microscopy study of Alzheimer'sbeta-amyloid peptide on different substrates: new insights into mechanism of beta-sheetformation. Proc Natl Acad Sci U S A.1999,96(7):3688-3693.
[37] Maltseva E, Kerth A, Blume A, et al. Adsorption of amyloid beta (1-40) peptide atphospholipid monolayers. Chembiochem.2005,6(10):1817-1824.
[38] Bush A I, Tanzi R E. Therapeutics for Alzheimer's disease based on the metal hypothesis.Neurotherapeutics.2008,5(3):421-432.
[39] Sohal R S. Oxidative stress hypothesis of aging. Free Radic Biol Med.2002,33(5):573-574.
[40] Boveris A, Oshino N, Chance B. The cellμlar production of hydrogen peroxide. Biochem J.1972,128(3):617-630.
[41] Andersen J K. Oxidative stress in neurodegeneration: cause or consequence? Nat Med.2004,10Suppl: S18-S25.
[42] Soragni A, Zambelli B, Mukrasch M D, et al. Structural characterization of binding of Cu(II)to tau protein. Biochemistry.2008,47(41):10841-10851.
[43]丰马庆.阿尔茨海默病相关多肽与Cu2+结合性质的研究[博士学位论文]北京：清华大学化学系.2006.
[44] Dias-Santagata D, Fulga T A, Duttaroy A, et al. Oxidative stress mediates tau-inducedneurodegeneration in Drosophila. J Clin Invest.2007,117(1):236-245.
[45] Lovell M A, Xiong S, Xie C, et al. Induction of hyperphosphorylated tau in primary ratcortical neuron cμltures mediated by oxidative stress and glycogen synthase kinase-3. JAlzheimers Dis.2004,6(6):659-671,673-681.
[46] Su X Y, Wu W H, Huang Z P, et al. Hydrogen peroxide can be generated by tau in thepresence of Cu(II). Biochem Biophys Res Commun.2007,358(2):661-665.
[47]苏小阳.铜离子催化Tau蛋白片段氧化及活性氧的产生北京：清华大学化学系.2007.
[48] Seo J, Lee K J. Post-translational modifications and their biological functions: proteomicanalysis and systematic approaches. J Biochem Mol Biol.2004,37(1):35-44.
[49] Ciechanover A, Brundin P. The ubiquitin proteasome system in neurodegenerative diseases:sometimes the chicken, sometimes the egg. Neuron.2003,40(2):427-446.
[50] Juan W C, Ong S T. The role of protein phosphorylation in therapy resistance and diseaseprogression in chronic myelogenous leukemia. Prog Mol Biol Transl Sci.2012,106:107-142.
[51] Ballatore C, Lee V M, Trojanowski J Q. Tau-mediated neurodegeneration in Alzheimer'sdisease and related disorders. Nat Rev Neurosci.2007,8(9):663-672.
[52] Liu F, Iqbal K, Grundke-Iqbal I, et al. O-GlcNAcylation regulates phosphorylation of tau: amechanism involved in Alzheimer's disease. Proc Natl Acad Sci U S A.2004,101(29):10804-10809.
[53] Martin L, Latypova X, Terro F. Post-translational modifications of tau protein: implicationsfor Alzheimer's disease. Neurochem Int.2011,58(4):458-471.
[54] Hatakeyama S, Matsumoto M, Kamura T, et al. U-box protein carboxyl terminus ofHsc70-interacting protein (CHIP) mediates poly-ubiquitylation preferentially on four-repeatTau and is involved in neurodegeneration of tauopathy. J Neurochem.2004,91(2):299-307.
[55] Okochi M, Walter J, Koyama A, et al. Constitutive phosphorylation of the Parkinson's diseaseassociated alpha-synuclein. J Biol Chem.2000,275(1):390-397.
[56] Chen L, Wei Y, Wang X, et al. Ribosylation rapidly induces alpha-synuclein to form highlycytotoxic molten globules of advanced glycation end products. PLoS One.2010,5(2): e9052.
[57] Tofaris G K, Kim H T, Hourez R, et al. Ubiquitin ligase Nedd4promotes alpha-synucleindegradation by the endosomal-lysosomal pathway. Proc Natl Acad Sci U S A.2011,108(41):17004-17009.
[58] Aiken C T, Steffan J S, Guerrero C M, et al. Phosphorylation of threonine3: implications forHuntingtin aggregation and neurotoxicity. J Biol Chem.2009,284(43):29427-29436.
[59] Wilcox K C, Zhou L, Jordon J K, et al. Modifications of superoxide dismutase (SOD1) inhuman erythrocytes: a possible role in amyotrophic lateral sclerosis. J Biol Chem.2009,284(20):13940-13947.
[60] Neumann M, Kwong L K, Lee E B, et al. Phosphorylation of S409/410of TDP-43is aconsistent feature in all sporadic and familial forms of TDP-43proteinopathies. ActaNeuropathol.2009,117(2):137-149.
[61] Buee L, Bussiere T, Buee-Scherrer V, et al. Tau protein isoforms, phosphorylation and role inneurodegenerative disorders. Brain Res Brain Res Rev.2000,33(1):95-130.
[62] Ballatore C, Lee V M, Trojanowski J Q. Tau-mediated neurodegeneration in Alzheimer'sdisease and related disorders. Nat Rev Neurosci.2007,8(9):663-672.
[63] Mandelkow E M A E. Tau in Alzheimer’s disease Trends Cell Biol.1998,8:425-427.
[64] Jin M, Shepardson N, Yang T, et al. Soluble amyloid beta-protein dimers isolated fromAlzheimer cortex directly induce Tau hyperphosphorylation and neuritic degeneration. ProcNatl Acad Sci U S A.2011,108(14):5819-5824.
[65] Kerscher O, Felberbaum R, Hochstrasser M. Modification of proteins by ubiquitin andubiquitin-like proteins. Annu Rev Cell Dev Biol.2006,22:159-180.
[66] Li W, Ye Y. Polyubiquitin chains: functions, structures, and mechanisms. Cell Mol Life Sci.2008,65(15):2397-2406.
[67] Kerscher O, Felberbaum R, Hochstrasser M. Modification of proteins by ubiquitin andubiquitin-like proteins. Annu Rev Cell Dev Biol.2006,22:159-180.
[68] Chatterjee C, Mcginty R K, Fierz B, et al. Disulfide-directed histone ubiquitylation revealsplasticity in hDot1L activation. Nat Chem Biol.2010,6(4):267-269.
[69] Muir T W, Sondhi D, Cole P A. Expressed protein ligation: a general method for proteinengineering. Proc Natl Acad Sci U S A.1998,95(12):6705-6710.
[70] Liu C C, Schμltz P G. Adding new chemistries to the genetic code. Annu Rev Biochem.2010,79:413-444.
[71] Wu Y W, Goody R S. Probing protein function by chemical modification. J Pept Sci.2010,16(10):514-523.
[72] Mcginty R K, Kim J, Chatterjee C, et al. Chemically ubiquitylated histone H2B stimulateshDot1L-mediated intranucleosomal methylation. Nature.2008,453(7196):812-816.
[73] Yang R, Pasunooti K K, Li F, et al. Synthesis of K48-linked diubiquitin using dual nativechemical ligation at lysine. Chem Commun (Camb).2010,46(38):7199-7201.
[74] Spasser L, Brik A. Chemistry and biology of the ubiquitin signal. Angew Chem Int Ed Engl.2012,51(28):6840-6862.
[75] Meier F, Abeywardana T, Dhall A, et al. Semisynthetic, site-specific ubiquitin modification ofalpha-synuclein reveals differential effects on aggregation. J Am Chem Soc.2012,134(12):5468-5471.
[76] Westermark P, Wernstedt C, Wilander E, et al. Amyloid fibrils in human insulinoma and isletsof Langerhans of the diabetic cat are derived from a neuropeptide-like protein also present innormal islet cells. Proc Natl Acad Sci U S A.1987,84(11):3881-3885.
[77] Cooper G J, Willis A C, Clark A, et al. Purification and characterization of a peptide fromamyloid-rich pancreases of type2diabetic patients. Proc Natl Acad Sci U S A.1987,84(23):8628-8632.
[78] Mishra R, Bμlic B, Sellin D, et al. Small-molecule inhibitors of islet amyloid polypeptide fibrilformation. Angew Chem Int Ed Engl.2008,47(25):4679-4682.
[79] Gilead S, Wolfenson H, Gazit E. Molecμlar mapping of the recognition interface between theislet amyloid polypeptide and insμlin. Angew Chem Int Ed Engl.2006,45(39):6476-6480.
[80] Knight J D, Williamson J A, Miranker A D. Interaction of membrane-bound islet amyloidpolypeptide with soluble and crystalline insulin. Protein Sci.2008,17(10):1850-1856.
[81] Zhang Y, Luo Y, Deng Y, et al. Lipid interaction and membrane perturbation of human isletamyloid polypeptide monomer and dimer by molecular dynamics simulations. PLoS One.2012,7(5): e38191.
[82] Knight J D, Hebda J A, Miranker A D. Conserved and cooperative assembly ofmembrane-bound alpha-helical states of islet amyloid polypeptide. Biochemistry.2006,45(31):9496-9508.
[83] Quist A, Doudevski I, Lin H, et al. Amyloid ion channels: a common structural link forprotein-misfolding disease. Proc Natl Acad Sci U S A.2005,102(30):10427-10432.
[84] Marchetti P, Bugliani M, Lupi R, et al. The endoplasmic reticulum in pancreatic beta cells oftype2diabetes patients. Diabetologia.2007,50(12):2486-2494.
[85] Ritzel R A, Meier J J, Lin C Y, et al. Human islet amyloid polypeptide oligomers disrupt cellcoupling, induce apoptosis, and impair insulin secretion in isolated human islets. Diabetes.2007,56(1):65-71.
[86] Ono K, Condron M M, Teplow D B. Structure-neurotoxicity relationships of amyloidbeta-protein oligomers. Proc Natl Acad Sci U S A.2009,106(35):14745-14750.
[87] Kayed R, Head E, Thompson J L, et al. Common structure of soluble amyloid oligomersimplies common mechanism of pathogenesis. Science.2003,300(5618):486-489.
[88] Dong M, Husale S, Sahin O. Determination of protein structural flexibility by microsecondforce spectroscopy. Nat Nanotechnol.2009,4(8):514-517.
[89] Dong M, Hovgaard M B, Mamdouh W, et al. AFM-based force spectroscopy measurements ofmature amyloid fibrils of the peptide glucagon. Nanotechnology.2008,19(38):384013.
[90] Dong M, Xu S, Oliveira C L, et al. Conformational changes in mannan-binding lectin bound toligand surfaces. J Immunol.2007,178(5):3016-3022.
[91] Howard K A, Dong M, Oupicky D, et al. Nanocarrier stimuli-activated gene delivery. Small.2007,3(1):54-57.
[92] Gan Y. Atomic and subnanometer resolution in ambient conditions by atomic forcemicroscopy Surface Science Reports64(2009)99-121.2008.
[93] Roach P, Farrar D, Perry C C. Interpretation of protein adsorption: surface-inducedconformational changes. J Am Chem Soc.2005,127(22):8168-8173.
[94] Keller C A, Kasemo B. Surface specific kinetics of lipid vesicle adsorption measured with aquartz crystal microbalance. Biophys J.1998,75(3):1397-1402.
[95] Dzwolak W, Grudzielanek S, Smirnovas V, et al. Ethanol-perturbed amyloidogenicself-assembly of insulin: looking for origins of amyloid strains. Biochemistry.2005,44(25):8948-8958.
[96] Jansen R, Grudzielanek S, Dzwolak W, et al. High pressure promotes circularly shaped insulinamyloid. J Mol Biol.2004,338(2):203-206.
[97] Barnham K J, Masters C L, Bush A I. Neurodegenerative diseases and oxidative stress. NatRev Drug Discov.2004,3(3):205-214.
[98] Christen Y. Oxidative stress and Alzheimer disease. Am J Clin Nutr.2000,71(2):621S-629S.
[99] Gaggelli E, Kozlowski H, Valensin D, et al. Copper homeostasis and neurodegenerativedisorders (Alzheimer's, prion, and Parkinson's diseases and amyotrophic lateral sclerosis).Chem Rev.2006,106(6):1995-2044.
[100] Valko M, Morris H, Cronin M T. Metals, toxicity and oxidative stress. Curr Med Chem.2005,12(10):1161-1208.
[101] Perry G, Sayre L M, Atwood C S, et al. The role of iron and copper in the aetiology ofneurodegenerative disorders: therapeutic implications. CNS Drugs.2002,16(5):339-352.
[102] Atwood C S, Huang X, Moir R D, et al. Role of free radicals and metal ions in thepathogenesis of Alzheimer's disease. Met Ions Biol Syst.1999,36:309-364.
[103] Huang X, Cuajungco M P, Atwood C S, et al. Cu(II) potentiation of alzheimer abetaneurotoxicity. Correlation with cell-free hydrogen peroxide production and metal reduction. JBiol Chem.1999,274(52):37111-37116.
[104] Huang X, Atwood C S, Hartshorn M A, et al. The A beta peptide of Alzheimer's diseasedirectly produces hydrogen peroxide through metal ion reduction. Biochemistry.1999,38(24):7609-7616.
[105] Huang X, Moir R D, Tanzi R E, et al. Redox-active metals, oxidative stress, and Alzheimer'sdisease pathology. Ann N Y Acad Sci.2004,1012:153-163.
[106] X.d. Huang R D M R. Redox-active metals, oxidative stress, and Alzheimer's diseasepathology, Redox-Active Metals in Neurological Disorders New York Acad Sciences, NewYork.2004:153-163.
[107] Bush A I. Copper, zinc, and the metallobiology of Alzheimer disease. Alzheimer Dis AssocDisord.2003,17(3):147-150.
[108] Sayre L M, Perry G, Harris P L, et al. In situ oxidative catalysis by neurofibrillary tangles andsenile plaques in Alzheimer's disease: a central role for bound transition metals. J Neurochem.2000,74(1):270-279.
[109] Grundke-Iqbal I, Iqbal K, Tung Y C, et al. Abnormal phosphorylation of themicrotubule-associated protein tau (tau) in Alzheimer cytoskeletal pathology. Proc Natl AcadSci U S A.1986,83(13):4913-4917.
[110] Grundke-Iqbal I, Iqbal K, Quinlan M, et al. Microtubule-associated protein tau. A componentof Alzheimer paired helical filaments. J Biol Chem.1986,261(13):6084-6089.
[111] G. Lee N C A M. The primary structure and heterogeneity of tau protein from mouse brain[Z].1998:239,285-288.
[112] Du Q.f. Ma Y M L J. Copper binding properties of a tau peptide associated with Alzheimer'sdisease studied by CD, NMR, and MALDI-TOF MS[Z].2006:27(),841-849.
[113] Du O.f. Ma Y M L J. Binding of copper (II) ion to an Alzheimer's tau peptide as revealed byMALDI-TOF MS, CD, and NMR. Biopolymers[Z].2005:79,74-85.
[114] Soragni A, Zambelli B, Mukrasch M D, et al. Structural characterization of binding of Cu(II)to tau protein. Biochemistry.2008,47(41):10841-10851.
[115] Su X Y, Wu W H, Huang Z P, et al. Hydrogen peroxide can be generated by tau in thepresence of Cu(II). Biochem Biophys Res Commun.2007,358(2):661-665.
[116] Ballatore C, Lee V M, Trojanowski J Q. Tau-mediated neurodegeneration in Alzheimer'sdisease and related disorders. Nat Rev Neurosci.2007,8(9):663-672.
[117] Mazanetz M P, Fischer P M. Untangling tau hyperphosphorylation in drug design forneurodegenerative diseases. Nat Rev Drug Discov.2007,6(6):464-479.
[118] Fields G B, Noble R L. Solid phase peptide synthesis utilizing9-fluorenylmethoxycarbonylamino acids. Int J Pept Protein Res.1990,35(3):161-214.
[119] Gomez L, Gellibert F, Wagner A, et al. An efficient procedure for traceless solid-phasesynthesis of N,N'-substituted thioureas by thermolytic cleavage of resin-bounddithiocarbamates. J Comb Chem.2000,2(1):75-79.
[120] Muhlradt P F, Kiess M, Meyer H, et al. Isolation, structure elucidation, and synthesis of amacrophage stimulatory lipopeptide from Mycoplasma fermentans acting at picomolarconcentration. J Exp Med.1997,185(11):1951-1958.
[121] G.f. Smith A D H W. New Colorimetric Reagent Specific for Cooper[Z].1953:25,510-511.
[122] Multhaup G, Schlicksupp A, Hesse L, et al. The amyloid precursor protein of Alzheimer'sdisease in the reduction of copper(II) to copper(I) Science.1996,271(5254):1406-1409.
[123] Ellman G L. Tissue sulfhydryl groups. Arch Biochem Biophys.1959,82(1):70-77.
[124] P.w. Riddles R L B A. ELLMANS REAGENT-5,5'-DITHIOBIS(2-NITROBENZOIC ACID)-RE-EXAMINATION[Z].1979:94,75-81.
[125] Manevich Y, Held K D, Biaglow J E. Coumarin-3-carboxylic acid as a detector for hydroxylradicals generated chemically and by gamma radiation. Radiat Res.1997,148(6):580-591.
[126] Liu L L, Franz K J. Phosphorylation of an alpha-synuclein peptide fragment enhances metalbinding. J Am Chem Soc.2005,127(27):9662-9663.
[127] Liu L L, Franz K J. Phosphorylation-dependent metal binding by alpha-synuclein peptidefragments. J Biol Inorg Chem.2007,12(2):234-247.
[128] Du Jt, Li Y M, Wei W, et al. Low-barrier hydrogen bond between phosphate and the amidegroup in phosphopeptide. J Am Chem Soc.2005,127(47):16350-16351.
[129] Kerscher O, Felberbaum R, Hochstrasser M. Modification of proteins by ubiquitin andubiquitin-like proteins. Annu Rev Cell Dev Biol.2006,22:159-180.
[130] Li W, Ye Y. Polyubiquitin chains: functions, structures, and mechanisms. Cell Mol Life Sci.2008,65(15):2397-2406.
[131] Mcginty R K, Chatterjee C, Muir T W. Semisynthesis of ubiquitylated proteins. MethodsEnzymol.2009,462:225-243.
[132] Chatterjee C, Mcginty R K, Pellois J P, et al. Auxiliary-mediated site-specific peptideubiquitylation. Angew Chem Int Ed Engl.2007,46(16):2814-2818.
[133] Mcginty R K, Kim J, Chatterjee C, et al. Chemically ubiquitylated histone H2B stimμlateshDot1L-mediated intranucleosomal methylation. Nature.2008,453(7196):812-816.
[134] Dawson P E, Muir T W, Clark-Lewis I, et al. Synthesis of proteins by native chemical ligation.Science.1994,266(5186):776-779.
[135] Saxon E, Bertozzi C R. Cell surface engineering by a modified Staudinger reaction. Science.2000,287(5460):2007-2010.
[136] Rostovtsev V V, Green L G, Fokin V V, et al. A stepwise huisgen cycloaddition process:copper(I)-catalyzed regioselective "ligation" of azides and terminal alkynes. Angew Chem IntEd Engl.2002,41(14):2596-2599.
[137] Sunbul M, Yin J. Site specific protein labeling by enzymatic posttranslational modification.Org Biomol Chem.2009,7(17):3361-3371.
[138] Liu C C, Schultz P G. Adding new chemistries to the genetic code. Annu Rev Biochem.2010,79:413-444.
[139] Young T S, Schultz P G. Beyond the canonical20amino acids: expanding the genetic lexicon.J Biol Chem.2010,285(15):11039-11044.
[140] Hejjaoui M, Haj-Yahya M, Kumar K S, et al. Towards elucidation of the role of ubiquitinationin the pathogenesis of Parkinson's disease with semisynthetic ubiquitinated alpha-synuclein.Angew Chem Int Ed Engl.2011,50(2):405-409.
[141]琳房川.阿尔兹海默病相关蛋白Aβ不同聚集体与铜离子的作用[硕士学位论文]北京：清华大学化学系.2010.
[142]刘磊，陈鹏，赵劲，何川.化学生物学基础[Z].科学出版社.
[2] Opie E L. The relation oe diabetes mellitus to lesions of the pancreas. hyaline degeneration ofthe islands oe langerhans. J Exp Med.1901,5(5):527-540.
[3] Soto C. Unfolding the role of protein misfolding in neurodegenerative diseases. Nat RevNeurosci.2003,4(1):49-60.
[4] Pardes H, Manton K G, Lander E S, et al. Effects of medical research on health care andeconomy. Science.1999,283(5398):36-37.
[5] Zimmet P. The burden of type2diabetes: are we doing enough? Diabetes Metab.2003,29(4Pt2):6S-9S.
[6] Zhang S, Liu H, Yu H, et al. Fas-associated death receptor signaling evoked by human amylinin islet beta-cells. Diabetes.2008,57(2):348-356.
[7] Hoppener J W, Lips C J. Role of islet amyloid in type2diabetes mellitus. Int J Biochem CellBiol.2006,38(5-6):726-736.
[8] Butler A E, Janson J, Bonner-Weir S, et al. Beta-cell deficit and increased beta-cell apoptosisin humans with type2diabetes. Diabetes.2003,52(1):102-110.
[9] Ward B, Walker K, Exley C. Copper(II) inhibits the formation of amylin amyloid in vitro. JInorg Biochem.2008,102(2):371-375.
[10] Sloane P D, Zimmerman S, Suchindran C, et al. The public health impact of Alzheimer'sdisease,2000-2050: potential implication of treatment advances. Annu Rev Public Health.2002,23:213-231.
[11] Mandelkow E. Alzheimer's disease. The tangled tale of tau. Nature.1999,402(6762):588-589.
[12] Gong C X, Liu F, Grundke-Iqbal I, et al. Post-translational modifications of tau protein inAlzheimer's disease.[J]. J Neural Transm.2005,112(6):813-838.
[13] Buee L, Bussiere T, Buee-Scherrer V, et al. Tau protein isoforms, phosphorylation and role inneurodegenerative disorders. Brain Res Brain Res Rev.2000,33(1):95-130.
[14] Dobson C M. The structural basis of protein folding and its links with human disease. PhilosTrans R Soc Lond B Biol Sci.2001,356(1406):133-145.
[15] Cohen F E, Pan K M, Huang Z, et al. Structural clues to prion replication.Science.1994,264(5158):530-531.
[16]陈媚莎.淀粉样蛋白聚集的机理研究和抑制剂的设计、合成与评价[博士学位论文].北京:清华大学化学系.2012.
[17] Salmona M, Malesani P, De Gioia L, et al. Molecμlar determinants of the physicochemicalproperties of a critical prion protein region comprising residues106-126. Biochem J.1999,342(Pt1):207-214.
[18] De Gioia L, Selvaggini C, Ghibaudi E, et al. Conformational polymorphism of theamyloidogenic and neurotoxic peptide homologous to residues106-126of the prion protein. JBiol Chem.1994,269(11):7859-7862.
[19] Jobling M F, Stewart L R, White A R, et al. The hydrophobic core sequence modμlates theneurotoxic and secondary structure properties of the prion peptide106-126. J Neurochem.1999,73(4):1557-1565.
[20] Dupiereux I, Zorzi W, Lins L, et al. Interaction of the106-126prion peptide with lipidmembranes and potential implication for neurotoxicity. Biochem Biophys Res Commun.2005,331(4):894-901.
[21] Kayed R, Head E, Thompson J L, et al. Common structure of soluble amyloid oligomersimplies common mechanism of pathogenesis. Science.2003,300(5618):486-489.
[22] Zraika S, Hμll R L, Udayasankar J, et al. Oxidative stress is induced by islet amyloidformation and time-dependently mediates amyloid-induced beta cell apoptosis. Diabetologia.2009,52(4):626-635.
[23] Janson J, Ashley R H, Harrison D, et al. The mechanism of islet amyloid polypeptide toxicityis membrane disruption by intermediate-sized toxic amyloid particles. Diabetes.1999,48(3):491-498.
[24] Mirzabekov T A, Lin M C, Kagan B L. Pore formation by the cytotoxic islet amyloid peptideamylin. J Biol Chem.1996,271(4):1988-1992.
[25] Brundin P, Melki R, Kopito R. Prion-like transmission of protein aggregates inneurodegenerative diseases. Nat Rev Mol Cell Biol.2010,11(4):301-307.
[26] Khemtemourian L, Killian J A, Hoppener J W, et al. Recent insights in islet amyloidpolypeptide-induced membrane disruption and its role in beta-cell death in type2diabetesmellitus. Exp Diabetes Res.2008,2008:421287.
[27] Kudva Y C, Mueske C, Butler P C, et al. A novel assay in vitro of human islet amyloidpolypeptide amyloidogenesis and effects of insμlin secretory vesicle peptides on amyloidformation. Biochem J.1998,331(Pt3):809-813.
[28] Janciauskiene S, Eriksson S, Carlemalm E, et al. B cell granμle peptides affect human isletamyloid polypeptide (IAPP) fibril formation in vitro. Biochem Biophys Res Commun.1997,236(3):580-585.
[29] Cui W, Ma J W, Lei P, et al. Insμlin is a kinetic but not a thermodynamic inhibitor of amylinaggregation. FEBS J.2009,276(12):3365-3371.
[30] Zraika S, Hμll R L, Udayasankar J, et al. Oxidative stress is induced by islet amyloidformation and time-dependently mediates amyloid-induced beta cell apoptosis. Diabetologia.2009,52(4):626-635.
[31] Masad A, Hayes L, Tabner B J, et al. Copper-mediated formation of hydrogen peroxide fromthe amylin peptide: a novel mechanism for degeneration of islet cells in type-2diabetesmellitus? FEBS Lett.2007,581(18):3489-3493.
[32] Butterfield S M, Lashuel H A. Amyloidogenic protein-membrane interactions: mechanisticinsight from model systems. Angew Chem Int Ed Engl.2010,49(33):5628-5654.
[33] Brender J R, Durr U H, Heyl D, et al. Membrane fragmentation by an amyloidogenic fragmentof human Islet Amyloid Polypeptide detected by solid-state NMR spectroscopy of membranenanotubes. Biochim Biophys Acta.2007,1768(9):2026-2029.
[34] Knight J D, Miranker A D. Phospholipid catalysis of diabetic amyloid assembly. J Mol Biol.2004,341(5):1175-1187.
[35] Ambroggio E E, Kim D H, Separovic F, et al. Surface behavior and lipid interaction ofAlzheimer beta-amyloid peptide1-42: a membrane-disrupting peptide. Biophys J.2005,88(4):2706-2713.
[36] Kowalewski T, Holtzman D M. In situ atomic force microscopy study of Alzheimer'sbeta-amyloid peptide on different substrates: new insights into mechanism of beta-sheetformation. Proc Natl Acad Sci U S A.1999,96(7):3688-3693.
[37] Maltseva E, Kerth A, Blume A, et al. Adsorption of amyloid beta (1-40) peptide atphospholipid monolayers. Chembiochem.2005,6(10):1817-1824.
[38] Bush A I, Tanzi R E. Therapeutics for Alzheimer's disease based on the metal hypothesis.Neurotherapeutics.2008,5(3):421-432.
[39] Sohal R S. Oxidative stress hypothesis of aging. Free Radic Biol Med.2002,33(5):573-574.
[40] Boveris A, Oshino N, Chance B. The cellμlar production of hydrogen peroxide. Biochem J.1972,128(3):617-630.
[41] Andersen J K. Oxidative stress in neurodegeneration: cause or consequence? Nat Med.2004,10Suppl: S18-S25.
[42] Soragni A, Zambelli B, Mukrasch M D, et al. Structural characterization of binding of Cu(II)to tau protein. Biochemistry.2008,47(41):10841-10851.
[43]丰马庆.阿尔茨海默病相关多肽与Cu2+结合性质的研究[博士学位论文]北京：清华大学化学系.2006.
[44] Dias-Santagata D, Fulga T A, Duttaroy A, et al. Oxidative stress mediates tau-inducedneurodegeneration in Drosophila. J Clin Invest.2007,117(1):236-245.
[45] Lovell M A, Xiong S, Xie C, et al. Induction of hyperphosphorylated tau in primary ratcortical neuron cμltures mediated by oxidative stress and glycogen synthase kinase-3. JAlzheimers Dis.2004,6(6):659-671,673-681.
[46] Su X Y, Wu W H, Huang Z P, et al. Hydrogen peroxide can be generated by tau in thepresence of Cu(II). Biochem Biophys Res Commun.2007,358(2):661-665.
[47]苏小阳.铜离子催化Tau蛋白片段氧化及活性氧的产生北京：清华大学化学系.2007.
[48] Seo J, Lee K J. Post-translational modifications and their biological functions: proteomicanalysis and systematic approaches. J Biochem Mol Biol.2004,37(1):35-44.
[49] Ciechanover A, Brundin P. The ubiquitin proteasome system in neurodegenerative diseases:sometimes the chicken, sometimes the egg. Neuron.2003,40(2):427-446.
[50] Juan W C, Ong S T. The role of protein phosphorylation in therapy resistance and diseaseprogression in chronic myelogenous leukemia. Prog Mol Biol Transl Sci.2012,106:107-142.
[51] Ballatore C, Lee V M, Trojanowski J Q. Tau-mediated neurodegeneration in Alzheimer'sdisease and related disorders. Nat Rev Neurosci.2007,8(9):663-672.
[52] Liu F, Iqbal K, Grundke-Iqbal I, et al. O-GlcNAcylation regulates phosphorylation of tau: amechanism involved in Alzheimer's disease. Proc Natl Acad Sci U S A.2004,101(29):10804-10809.
[53] Martin L, Latypova X, Terro F. Post-translational modifications of tau protein: implicationsfor Alzheimer's disease. Neurochem Int.2011,58(4):458-471.
[54] Hatakeyama S, Matsumoto M, Kamura T, et al. U-box protein carboxyl terminus ofHsc70-interacting protein (CHIP) mediates poly-ubiquitylation preferentially on four-repeatTau and is involved in neurodegeneration of tauopathy. J Neurochem.2004,91(2):299-307.
[55] Okochi M, Walter J, Koyama A, et al. Constitutive phosphorylation of the Parkinson's diseaseassociated alpha-synuclein. J Biol Chem.2000,275(1):390-397.
[56] Chen L, Wei Y, Wang X, et al. Ribosylation rapidly induces alpha-synuclein to form highlycytotoxic molten globules of advanced glycation end products. PLoS One.2010,5(2): e9052.
[57] Tofaris G K, Kim H T, Hourez R, et al. Ubiquitin ligase Nedd4promotes alpha-synucleindegradation by the endosomal-lysosomal pathway. Proc Natl Acad Sci U S A.2011,108(41):17004-17009.
[58] Aiken C T, Steffan J S, Guerrero C M, et al. Phosphorylation of threonine3: implications forHuntingtin aggregation and neurotoxicity. J Biol Chem.2009,284(43):29427-29436.
[59] Wilcox K C, Zhou L, Jordon J K, et al. Modifications of superoxide dismutase (SOD1) inhuman erythrocytes: a possible role in amyotrophic lateral sclerosis. J Biol Chem.2009,284(20):13940-13947.
[60] Neumann M, Kwong L K, Lee E B, et al. Phosphorylation of S409/410of TDP-43is aconsistent feature in all sporadic and familial forms of TDP-43proteinopathies. ActaNeuropathol.2009,117(2):137-149.
[61] Buee L, Bussiere T, Buee-Scherrer V, et al. Tau protein isoforms, phosphorylation and role inneurodegenerative disorders. Brain Res Brain Res Rev.2000,33(1):95-130.
[62] Ballatore C, Lee V M, Trojanowski J Q. Tau-mediated neurodegeneration in Alzheimer'sdisease and related disorders. Nat Rev Neurosci.2007,8(9):663-672.
[63] Mandelkow E M A E. Tau in Alzheimer’s disease Trends Cell Biol.1998,8:425-427.
[64] Jin M, Shepardson N, Yang T, et al. Soluble amyloid beta-protein dimers isolated fromAlzheimer cortex directly induce Tau hyperphosphorylation and neuritic degeneration. ProcNatl Acad Sci U S A.2011,108(14):5819-5824.
[65] Kerscher O, Felberbaum R, Hochstrasser M. Modification of proteins by ubiquitin andubiquitin-like proteins. Annu Rev Cell Dev Biol.2006,22:159-180.
[66] Li W, Ye Y. Polyubiquitin chains: functions, structures, and mechanisms. Cell Mol Life Sci.2008,65(15):2397-2406.
[67] Kerscher O, Felberbaum R, Hochstrasser M. Modification of proteins by ubiquitin andubiquitin-like proteins. Annu Rev Cell Dev Biol.2006,22:159-180.
[68] Chatterjee C, Mcginty R K, Fierz B, et al. Disulfide-directed histone ubiquitylation revealsplasticity in hDot1L activation. Nat Chem Biol.2010,6(4):267-269.
[69] Muir T W, Sondhi D, Cole P A. Expressed protein ligation: a general method for proteinengineering. Proc Natl Acad Sci U S A.1998,95(12):6705-6710.
[70] Liu C C, Schμltz P G. Adding new chemistries to the genetic code. Annu Rev Biochem.2010,79:413-444.
[71] Wu Y W, Goody R S. Probing protein function by chemical modification. J Pept Sci.2010,16(10):514-523.
[72] Mcginty R K, Kim J, Chatterjee C, et al. Chemically ubiquitylated histone H2B stimulateshDot1L-mediated intranucleosomal methylation. Nature.2008,453(7196):812-816.
[73] Yang R, Pasunooti K K, Li F, et al. Synthesis of K48-linked diubiquitin using dual nativechemical ligation at lysine. Chem Commun (Camb).2010,46(38):7199-7201.
[74] Spasser L, Brik A. Chemistry and biology of the ubiquitin signal. Angew Chem Int Ed Engl.2012,51(28):6840-6862.
[75] Meier F, Abeywardana T, Dhall A, et al. Semisynthetic, site-specific ubiquitin modification ofalpha-synuclein reveals differential effects on aggregation. J Am Chem Soc.2012,134(12):5468-5471.
[76] Westermark P, Wernstedt C, Wilander E, et al. Amyloid fibrils in human insulinoma and isletsof Langerhans of the diabetic cat are derived from a neuropeptide-like protein also present innormal islet cells. Proc Natl Acad Sci U S A.1987,84(11):3881-3885.
[77] Cooper G J, Willis A C, Clark A, et al. Purification and characterization of a peptide fromamyloid-rich pancreases of type2diabetic patients. Proc Natl Acad Sci U S A.1987,84(23):8628-8632.
[78] Mishra R, Bμlic B, Sellin D, et al. Small-molecule inhibitors of islet amyloid polypeptide fibrilformation. Angew Chem Int Ed Engl.2008,47(25):4679-4682.
[79] Gilead S, Wolfenson H, Gazit E. Molecμlar mapping of the recognition interface between theislet amyloid polypeptide and insμlin. Angew Chem Int Ed Engl.2006,45(39):6476-6480.
[80] Knight J D, Williamson J A, Miranker A D. Interaction of membrane-bound islet amyloidpolypeptide with soluble and crystalline insulin. Protein Sci.2008,17(10):1850-1856.
[81] Zhang Y, Luo Y, Deng Y, et al. Lipid interaction and membrane perturbation of human isletamyloid polypeptide monomer and dimer by molecular dynamics simulations. PLoS One.2012,7(5): e38191.
[82] Knight J D, Hebda J A, Miranker A D. Conserved and cooperative assembly ofmembrane-bound alpha-helical states of islet amyloid polypeptide. Biochemistry.2006,45(31):9496-9508.
[83] Quist A, Doudevski I, Lin H, et al. Amyloid ion channels: a common structural link forprotein-misfolding disease. Proc Natl Acad Sci U S A.2005,102(30):10427-10432.
[84] Marchetti P, Bugliani M, Lupi R, et al. The endoplasmic reticulum in pancreatic beta cells oftype2diabetes patients. Diabetologia.2007,50(12):2486-2494.
[85] Ritzel R A, Meier J J, Lin C Y, et al. Human islet amyloid polypeptide oligomers disrupt cellcoupling, induce apoptosis, and impair insulin secretion in isolated human islets. Diabetes.2007,56(1):65-71.
[86] Ono K, Condron M M, Teplow D B. Structure-neurotoxicity relationships of amyloidbeta-protein oligomers. Proc Natl Acad Sci U S A.2009,106(35):14745-14750.
[87] Kayed R, Head E, Thompson J L, et al. Common structure of soluble amyloid oligomersimplies common mechanism of pathogenesis. Science.2003,300(5618):486-489.
[88] Dong M, Husale S, Sahin O. Determination of protein structural flexibility by microsecondforce spectroscopy. Nat Nanotechnol.2009,4(8):514-517.
[89] Dong M, Hovgaard M B, Mamdouh W, et al. AFM-based force spectroscopy measurements ofmature amyloid fibrils of the peptide glucagon. Nanotechnology.2008,19(38):384013.
[90] Dong M, Xu S, Oliveira C L, et al. Conformational changes in mannan-binding lectin bound toligand surfaces. J Immunol.2007,178(5):3016-3022.
[91] Howard K A, Dong M, Oupicky D, et al. Nanocarrier stimuli-activated gene delivery. Small.2007,3(1):54-57.
[92] Gan Y. Atomic and subnanometer resolution in ambient conditions by atomic forcemicroscopy Surface Science Reports64(2009)99-121.2008.
[93] Roach P, Farrar D, Perry C C. Interpretation of protein adsorption: surface-inducedconformational changes. J Am Chem Soc.2005,127(22):8168-8173.
[94] Keller C A, Kasemo B. Surface specific kinetics of lipid vesicle adsorption measured with aquartz crystal microbalance. Biophys J.1998,75(3):1397-1402.
[95] Dzwolak W, Grudzielanek S, Smirnovas V, et al. Ethanol-perturbed amyloidogenicself-assembly of insulin: looking for origins of amyloid strains. Biochemistry.2005,44(25):8948-8958.
[96] Jansen R, Grudzielanek S, Dzwolak W, et al. High pressure promotes circularly shaped insulinamyloid. J Mol Biol.2004,338(2):203-206.
[97] Barnham K J, Masters C L, Bush A I. Neurodegenerative diseases and oxidative stress. NatRev Drug Discov.2004,3(3):205-214.
[98] Christen Y. Oxidative stress and Alzheimer disease. Am J Clin Nutr.2000,71(2):621S-629S.
[99] Gaggelli E, Kozlowski H, Valensin D, et al. Copper homeostasis and neurodegenerativedisorders (Alzheimer's, prion, and Parkinson's diseases and amyotrophic lateral sclerosis).Chem Rev.2006,106(6):1995-2044.
[100] Valko M, Morris H, Cronin M T. Metals, toxicity and oxidative stress. Curr Med Chem.2005,12(10):1161-1208.
[101] Perry G, Sayre L M, Atwood C S, et al. The role of iron and copper in the aetiology ofneurodegenerative disorders: therapeutic implications. CNS Drugs.2002,16(5):339-352.
[102] Atwood C S, Huang X, Moir R D, et al. Role of free radicals and metal ions in thepathogenesis of Alzheimer's disease. Met Ions Biol Syst.1999,36:309-364.
[103] Huang X, Cuajungco M P, Atwood C S, et al. Cu(II) potentiation of alzheimer abetaneurotoxicity. Correlation with cell-free hydrogen peroxide production and metal reduction. JBiol Chem.1999,274(52):37111-37116.
[104] Huang X, Atwood C S, Hartshorn M A, et al. The A beta peptide of Alzheimer's diseasedirectly produces hydrogen peroxide through metal ion reduction. Biochemistry.1999,38(24):7609-7616.
[105] Huang X, Moir R D, Tanzi R E, et al. Redox-active metals, oxidative stress, and Alzheimer'sdisease pathology. Ann N Y Acad Sci.2004,1012:153-163.
[106] X.d. Huang R D M R. Redox-active metals, oxidative stress, and Alzheimer's diseasepathology, Redox-Active Metals in Neurological Disorders New York Acad Sciences, NewYork.2004:153-163.
[107] Bush A I. Copper, zinc, and the metallobiology of Alzheimer disease. Alzheimer Dis AssocDisord.2003,17(3):147-150.
[108] Sayre L M, Perry G, Harris P L, et al. In situ oxidative catalysis by neurofibrillary tangles andsenile plaques in Alzheimer's disease: a central role for bound transition metals. J Neurochem.2000,74(1):270-279.
[109] Grundke-Iqbal I, Iqbal K, Tung Y C, et al. Abnormal phosphorylation of themicrotubule-associated protein tau (tau) in Alzheimer cytoskeletal pathology. Proc Natl AcadSci U S A.1986,83(13):4913-4917.
[110] Grundke-Iqbal I, Iqbal K, Quinlan M, et al. Microtubule-associated protein tau. A componentof Alzheimer paired helical filaments. J Biol Chem.1986,261(13):6084-6089.
[111] G. Lee N C A M. The primary structure and heterogeneity of tau protein from mouse brain[Z].1998:239,285-288.
[112] Du Q.f. Ma Y M L J. Copper binding properties of a tau peptide associated with Alzheimer'sdisease studied by CD, NMR, and MALDI-TOF MS[Z].2006:27(),841-849.
[113] Du O.f. Ma Y M L J. Binding of copper (II) ion to an Alzheimer's tau peptide as revealed byMALDI-TOF MS, CD, and NMR. Biopolymers[Z].2005:79,74-85.
[114] Soragni A, Zambelli B, Mukrasch M D, et al. Structural characterization of binding of Cu(II)to tau protein. Biochemistry.2008,47(41):10841-10851.
[115] Su X Y, Wu W H, Huang Z P, et al. Hydrogen peroxide can be generated by tau in thepresence of Cu(II). Biochem Biophys Res Commun.2007,358(2):661-665.
[116] Ballatore C, Lee V M, Trojanowski J Q. Tau-mediated neurodegeneration in Alzheimer'sdisease and related disorders. Nat Rev Neurosci.2007,8(9):663-672.
[117] Mazanetz M P, Fischer P M. Untangling tau hyperphosphorylation in drug design forneurodegenerative diseases. Nat Rev Drug Discov.2007,6(6):464-479.
[118] Fields G B, Noble R L. Solid phase peptide synthesis utilizing9-fluorenylmethoxycarbonylamino acids. Int J Pept Protein Res.1990,35(3):161-214.
[119] Gomez L, Gellibert F, Wagner A, et al. An efficient procedure for traceless solid-phasesynthesis of N,N'-substituted thioureas by thermolytic cleavage of resin-bounddithiocarbamates. J Comb Chem.2000,2(1):75-79.
[120] Muhlradt P F, Kiess M, Meyer H, et al. Isolation, structure elucidation, and synthesis of amacrophage stimulatory lipopeptide from Mycoplasma fermentans acting at picomolarconcentration. J Exp Med.1997,185(11):1951-1958.
[121] G.f. Smith A D H W. New Colorimetric Reagent Specific for Cooper[Z].1953:25,510-511.
[122] Multhaup G, Schlicksupp A, Hesse L, et al. The amyloid precursor protein of Alzheimer'sdisease in the reduction of copper(II) to copper(I) Science.1996,271(5254):1406-1409.
[123] Ellman G L. Tissue sulfhydryl groups. Arch Biochem Biophys.1959,82(1):70-77.
[124] P.w. Riddles R L B A. ELLMANS REAGENT-5,5'-DITHIOBIS(2-NITROBENZOIC ACID)-RE-EXAMINATION[Z].1979:94,75-81.
[125] Manevich Y, Held K D, Biaglow J E. Coumarin-3-carboxylic acid as a detector for hydroxylradicals generated chemically and by gamma radiation. Radiat Res.1997,148(6):580-591.
[126] Liu L L, Franz K J. Phosphorylation of an alpha-synuclein peptide fragment enhances metalbinding. J Am Chem Soc.2005,127(27):9662-9663.
[127] Liu L L, Franz K J. Phosphorylation-dependent metal binding by alpha-synuclein peptidefragments. J Biol Inorg Chem.2007,12(2):234-247.
[128] Du Jt, Li Y M, Wei W, et al. Low-barrier hydrogen bond between phosphate and the amidegroup in phosphopeptide. J Am Chem Soc.2005,127(47):16350-16351.
[129] Kerscher O, Felberbaum R, Hochstrasser M. Modification of proteins by ubiquitin andubiquitin-like proteins. Annu Rev Cell Dev Biol.2006,22:159-180.
[130] Li W, Ye Y. Polyubiquitin chains: functions, structures, and mechanisms. Cell Mol Life Sci.2008,65(15):2397-2406.
[131] Mcginty R K, Chatterjee C, Muir T W. Semisynthesis of ubiquitylated proteins. MethodsEnzymol.2009,462:225-243.
[132] Chatterjee C, Mcginty R K, Pellois J P, et al. Auxiliary-mediated site-specific peptideubiquitylation. Angew Chem Int Ed Engl.2007,46(16):2814-2818.
[133] Mcginty R K, Kim J, Chatterjee C, et al. Chemically ubiquitylated histone H2B stimμlateshDot1L-mediated intranucleosomal methylation. Nature.2008,453(7196):812-816.
[134] Dawson P E, Muir T W, Clark-Lewis I, et al. Synthesis of proteins by native chemical ligation.Science.1994,266(5186):776-779.
[135] Saxon E, Bertozzi C R. Cell surface engineering by a modified Staudinger reaction. Science.2000,287(5460):2007-2010.
[136] Rostovtsev V V, Green L G, Fokin V V, et al. A stepwise huisgen cycloaddition process:copper(I)-catalyzed regioselective "ligation" of azides and terminal alkynes. Angew Chem IntEd Engl.2002,41(14):2596-2599.
[137] Sunbul M, Yin J. Site specific protein labeling by enzymatic posttranslational modification.Org Biomol Chem.2009,7(17):3361-3371.
[138] Liu C C, Schultz P G. Adding new chemistries to the genetic code. Annu Rev Biochem.2010,79:413-444.
[139] Young T S, Schultz P G. Beyond the canonical20amino acids: expanding the genetic lexicon.J Biol Chem.2010,285(15):11039-11044.
[140] Hejjaoui M, Haj-Yahya M, Kumar K S, et al. Towards elucidation of the role of ubiquitinationin the pathogenesis of Parkinson's disease with semisynthetic ubiquitinated alpha-synuclein.Angew Chem Int Ed Engl.2011,50(2):405-409.
[141]琳房川.阿尔兹海默病相关蛋白Aβ不同聚集体与铜离子的作用[硕士学位论文]北京：清华大学化学系.2010.
[142]刘磊，陈鹏，赵劲，何川.化学生物学基础[Z].科学出版社.