Amyloid Fibril Formation and Disaggregation of Fragment 1-29 of Apomyoglobin: Insights into the Effect of pH on Protein Fibrillogenesis

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• 作者：Paola Picott ; Giorgia De Franceschi ; Erica Frare ; Barbara Spolaore ; Marcello Zambonin ; Fabrizio Chiti ; Patrizia Polverino de Laureto ; Angelo Fontana
• 关键词：protein aggregation ; amyloid ; protein fragments ; electron microscopy ; circular dichroism
• 刊名：Journal of Molecular Biology
• 出版年：2007
• 出版时间：13 April 2007
• 年：2007
• 卷：367
• 期：5
• 页码：1237-1245
• 全文大小：1124 K

文摘

The N-terminal fragment 1–29 of horse heart apomyoglobin (apoMb_1–29) is highly prone to form amyloid-like fibrils at low pH. Fibrillogenesis at pH 2.0 occurs following a nucleation-dependent growth mechanism, as evidenced by the thioflavin T (ThT) assay. Transmission electron microscopy (TEM) confirms the presence of regular amyloid-like fibrils and far-UV circular dichroism (CD) spectra indicate the acquisition of a high content of β-sheet structure. ThT assay, TEM and CD highlight fast and complete disaggregation of the fibrils, if the pH of a suspension of mature fibrils is increased to 8.3. It is of interest that amyloid-like fibrils form again if the pH of the solution is brought back to 2.0. While apoMb_1–29 fibrils obtained at pH 2.0 are resistant to proteolysis by pepsin, the disaggregated fibrils are easily cleaved at pH 8.3 by trypsin and V8 protease, and some of the resulting fragments aggregate very quickly in the proteolysis mixture, forming amyloid-like fibrils. We show that the increase of amyloidogenicity of apoMb_1–29 following acidification or proteolysis at pH 8.3 can be attributed to the decrease of the peptide net charge following these alterations. The results observed here for apoMb_1–29 provide an experimental basis for explaining the effect of charge and pH on amyloid fibril formation by both unfolded and folded protein systems.