| Abstract:Alzheimer’s disease (AD) is a devastating neurodegenerative disease and the main etiology of AD is senile plaques (SP) and neurofibrillary tangles. Amyloid beta peptide (Aβ) is the main component of senile plaques. The hydrophobic core residues16-20(KLVFF) of amyloid beta also known as Tjernberg peptide is crucial for the formation of beta structures by amyloid beta. The pentapeptide KLVFF (Tjernberg peptide) has been reported to possess amyloid plaque blocking property. However, stability in plasma, water solubility and proteolytic stability of were poor limit it’s using as an Aβinhibitor. Many works have done to improve its properties. In this work, we designed and synthesized of a series of novel N-ferrocenoyl peptides by liquid-phase or solid-phase peptide synthesis method asβ-amyloid fibril assembly inhibitors.(1) Electrochemical peptide Fc-KLVFF(Fc:ferrocenoyl) and control peptide Boc-KLVFF were synthesized from ferrocene, Lys Leu, Val and Phe by liquid-phase peptide synthesis method using O-(benzotriazol-yl)-N,N,N’,N’-tetramethyluronium (HBTU)/1-hydroxybenzotrizole (HOBt) as coupling reagents, and then characterized by IR, UV, NMR and MS. The electrochemical results showed that Fc-KLVFF exhibits a pair of well-defined redox wave at oxidation potential (Epa) and reduction potential (Epc) at0.626V and0.544V, respectively, in the range of0.4-0.8V, and the ratio of Ipa/Ipc is1.08. Protease degradation of peptide derivatives Fc-KLVFF and Boc-KLVFF were evaluated used trypsin and chymotrypsin in vitro, and the degraded fragments were analyzed by HPLC. The enzymatic degradation experimental results showed that modification with ferrocene group, Fc-KLVFF showed increased stability as compared with Boc-KLVFF. It is in agreement with the experimental hypothesis; the modification with ferrocene increases the resistance of the compound to proteolysis. The partition coefficient log P of Fc-KLVFF and Boc-KLVFF were determined by RP-HPLC. The lipophilic evaluation studies found that they have a strong hydrophobic and not readily soluble in aqueous solution. Especially modification with ferrocene group, Fc-KLVFF hydrophobic stronger than KLVFF, because they the solubility in water is too small, so can not be directly applied to an aqueous solution.We used electrochemical properties of ferrocene as a rapid and effective determination of the interaction between Aβ342and Fc-KLVFF. The inhibitory effect on Aβ42. was also investigated using thioflavin T (ThT) fluorescence probe, atomic force microscopy (AFM) and transmission electron microscopy (TEM). The experimental results showed that they are inhibited the Aβ self-assembly into fibrils, but the inhibitory effect of Fc-KLVFF is better than that of Boc-KLVFF. We postulated that ferrocene may insert into (3-sheet structures and inhibit Aβ42fibrillization by its lipophilicity and space steric hindrance.(2) We employed Fmoc solid-phase synthesis method synthesized water-soluble better peptides, Ac-LPFFD and an N-ferrocenoyl peptide Fc-KLVFFK6as inhibitors of fibrillogenesis, which contained the hydrophobic sequence of Aβ16-20, could interact with the corresponding residues of A(3via self-cognition and inhibited the A(3self-assembly into fibrils. Thioflavin T (ThT) fluorescence spectroscopy and AFM were employed to investigate the interaction between inhibitors and Aβ42in vitro. The results revealed Fc-KLVFFK6inhibits A(3aggregation well.(3) The self-assembly of ferrocene modified hydrophobic pentapeptide fragment of (3-amyloid, Fc-KLVFF, is studied. Fc-KLVFF can self-assemble into nanotubes in methanol, as confirmed by transmission electron microscopy. The secondary structure of the nanotubes is characterized by FT-IR, CD, fluorescence spectroscopy and UV/visible spectroscopy, which indicated the important role of aromatic π-π stacking interactions between phenylalanine residues in driving a-helix and (3-sheet self-assembly. |
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