Research On The Mechanism And Regulation Of Amyloid Protein In Vivo

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.