Using bioinformatics methods of investigation, amyloidogenic regions of Aβ16-25, Aβ31-40, and Aβ33-42, responsible for the formation of fibrils, were determined in the Aβ1-42 peptide. Fragments were synthesized in sufficient quantities, and the quality of the preparations was verified by mass spectrometric analysis. To study their amyloidogenic properties, methods of fluorescence spectroscopy (ThT binding), electron microscopy, and X-ray diffraction analysis were used. All three fragments under conditions of 5% DMSO and 50 mM Tris-HCl (pH 7.5) interact with ThT. At the same time fragments of Aβ31-40 and Aβ33-42 polymerize in the form of fibrils of different morphology, and fragment Aβ16-25 form polymers that are not typical of most amyloids in the form of films. According to X-ray diffraction data, the fibrils of all fragments show the presence of two main reflexes (4.6-4.8 Ǻ and 8-12 Ǻ), characteristic of the cross-β structure, indicating their amyloid structure. Analysis of the experimental data and structural modeling allowed us to conclude that the main building block in the formation of fibrils by fragments of Aβ1-42 peptide is a ring oligomer. The interaction of ring oligomers in different ways leads to the formation of polymers of different morphologies. Based on the data set, a new model of fibrillation was proposed.
β1-42 peptide, amyloids, bioinformatics, new model of fibrillation
1. Surin A.K., Grigorashvili E.I., Suvorina M.Yu., Selivanova O.M., Galzitskaya O.V. Opredelenie uchastkov peptida Aβ(1-40), vovlechennyh v obrazovanie amiloidnyh fibrill. Biohimiya (Moskva), 2016, t. 81, s. 999-1007. [Surin A.K., Grigorashvili E.I., Suvorina M.Yu., Selivanova O.M., Galzitskaya O.V. Determination of regions involved in amyloid fibril formation for Aβ(1-40) peptide. Biochemistry (Moscow), 2016, vol. 81, pp. 762-769 (in Rus.)]
2. Suvorina M.Yu, Selivanova O.M, Grigorashvili E.I., Nikulin A.D, Marchenkov V.V., Surin A.K., Galzitskaya O.V. Studies of Polymorphism of Amyloid-β42Peptide from Different Suppliers. J. Alzheimer’s Dis., 2015, vol. 47, pp. 583-593.
3. Garbuzynskiy S.O., Lobanov M.Y., Galzitskaya O.V. FoldAmyloid: a method of prediction of amyloidogenic regions from protein sequence. Bioinformatics, 2010, vol. 26, pp.326-332.
4. Selivanova O.M., Surin A.K., Marchenkov V.V., Dzhus U.F., Grigorashvili E.I., Suvorina M.Yu., Glyakina A.V., Dovidchenko N.V., Galzitskaya O.V. The mechanism underlying amyloid polymorphism is opened for alzheimer’s disease amyloid-β peptide. J. Alzheimer Dis., 2016, vol. 54, pp. 821-830.
5. Makin O.S., Serpell L.C. X-ray diffraction studies of amyloid structure. Methods Mol. Biol., 2005, vol. 299, pp. 67-80.
6. Malinchik S.B., Inouye H., Szumowski K.E., Kirschner D.A. Structural Analysis of Alzheimer’s β(1- 40) Amyloid: Protofilament Assembly of Tubular Fibrils. Biophysical J., 1998, vol., pp. 537-545.
7. Selivanova O.M., Grigorashvili E.I., Suvirina M.Yu., Dzhus U.F., Nikulin A.D., Marchenkov V.V., Surin A.K., Glyakina A.V., Galzitskaya O.V. X-ray diffraction and electron microscopy data for amyloid formation of Aβ40 and Aβ42. Data in Brief, 2016, vol. 8, pp. 108-113.
8. Inouye H., Fraser P.E., Kirschner D.A. Structure of aB-crystallite assemblies formed by Alzheimer β-amyloid protein analogues: analysis by x-ray diffraction. Biophysical J., 1993, vol. 64, pp. 502-519.
9. Astbury W.T., Dickinson S., Bailey K. The X-ray interpretation of denaturation and the structure of the seed globulins. Biochem J., 1935, vol. 29, pp. 2351-2360.
10. Eanes E.D., Glenner G.G. X-ray diffraction studies on amyloid filaments. Histochem. Cytochem., 1968, vol. 16, pp. 673-677.
11. Cohen A.S., Calkins E. Electron microscopic observations on a fibrous component in amyloid of diverse origins. Nature, 1959, vol. 183, pp. 1202-1203.
12. Shirahama T., Cohen A.S. High-resolution electron microscopic analysis of the amyloid fibril. J. Cell. Biol,. 1967, vol. 33, pp. 679-708.
13. Benditt E.P., Eriksen N. Amyloid, III. A protein related to the subunit structure of human amyloid fibrils. Proc. Natl. Acad. Sci. USA, 1967, vol. 55, pp. 308-316.
14. Nielsen E.H., Nybo M., Svehag S.-E. Electron microscopy of prefibrillar structures and amyloid fibrils. Methods Enzymol., 1999, vol. 309, pp. 491-496.
15. Bhak G., Lee G-H., Hahn G-S., Paik S.R. Granular Assembly of α-Synuclein Leading to the Accelerated Amyloid Fibril Formation with Shear Stress. PLoS ONE, 2009, vol. 4, e4177.
16. Shankar G.M., Li S., Mehta T.H., Garcia-Munoz A., Shepardson N., Smith I., Brett F.M., Farell M.A., Rowan M.J., Lemere C.A., Regan C.M., Walsh D.M., Sabatini B.L., Selkoe D.J. Amyloid-β protein dimers isolated directly from alzheimer' brains impair synaptic plasticity and memory. Nature Medicine, 2008, vol. 14, pp. 837-842.
17. Quist A., Doudevski I., Lin H., Azimova R., Ng D., Frangione B., Kagan B., Ghiso J., Lal R. Amyloid ion channels: a common structural link for protein-misfolding disease. Proc. Natl. Acad. Sci. USA. 2005, vol. 102, pp. 10427-10432.
