Characterization of Chimaeras of a Thermoresponsive Polymer and the Parathyroid Hormone

Bruno Voigt, Zhanna Evgrafova, Monika Baumann, Madlen Stephani, Marvin Umlandt, Wolfgang H. Binder, Jochen Balbach

Faculty of Natural Sciences II, Martin-Luther-Universität Halle-Wittenberg, D-06120 Halle, Germany

The parathyroid hormone (PTH) is an 84 residue peptide from the parathyroid glands which controls the calcium and phosphate level in human blood. The peptide adopts an α-helical conformation at the N-terminus and is intrinsically disordered at the C-terminus. Amyloidogenic properties of PTH have been reported [1]. Here we show the in vitro formation of amyloid fibrils of PTH 1-84 and of the pharmaceutically relevant N-terminal fragment PTH 1-34 under physiological conditions. To get further insights into the mechanism of amyloid fibrillation we investigated the effect of thermoresponsive polymers [2] on PTH. We covalently attached polyacrylate based polymers to 15N isotope labelled PTH 1-84 and used two dimensional NMR techniques for the characterization of the resulting chimaeras. This allows the observation of amino acid sequence specific changes of the cross peaks corresponding to the peptide backbone according to the polymer state. The studies revealed strong dependencies of chemical shifts on the temperature, the peptide attachment site and the polymer molecular weight.

References
[1] Gopalswamy, M.; Kumar, A.; Adler, J.; Baumann, M.; Henze, M.; Kumar, S.T.; Fändrich, M.; Scheidt, H.A.; Huster, D.; Balbach, J., Biochim Biophys Acta 2015, 1854, 249-257. (link)
[2] Funtan, S.; Evgrafova, Z.; Adler, J.; Huster, D.; Binder, W.H., Polymers 2016, 8, 178. (link)

Amyloid Protein Aggregation in the Presence of Temperature-Sensitive Polymers

Zhanna Evgrafova1, Sonu Kumar1, Bruno Voigt1,  Juliane Adler2, Daniel Huster2, Jochen Balbach1 and Wolfgang H. Binder1

1Faculty of Natural Science II, Martin Luther University Halle-Wittenberg, Von-Danckelmann-Platz 4, D-06120 Halle (Saale), Germany
2Institute for Medical Physics and Biophysics, Leipzig University, Härtelstraße 16-18, D-04107 Leipzig, Germany

The formation of amyloid fibrils is considered to be one of the main causes for many neurodegenerative diseases, such as Alzheimer’s, Parkinson’s or Huntington’s disease [1, 2]. Current knowledge suggests that amyloid-aggregation represents a nucleation-dependent aggregation process in vitro, where a sigmoidal growth phase follows an induction period. Here, we studied the fibrillation of amyloid β 1-40 (Aβ40) and Parathyroid hormone (PTH) in the presence of thermoresponsive polymers, expected to alter their fibrillation kinetics due to their specific hydrophobic and hydrophilic interactions with proteins [3]. Mixtures in varying concentrations and the conjugates of PTH or Aβ40 with poly(ethylene glycol) methyl ether acrylate were studied via time-dependent measurements of the thioflavin T (ThT) fluorescence and transition electron microscopy (TEM). The studies revealed that amyloid fibrillation was altered, accompanied by either reduction or elongation of the lag phase of PTH or Aβ40 fibrillation in the presence of studied polymers [4].

References:
[1] Chiti, F.; Dobson, C.M., Annu. Rev. Biochem. 2006, 75, 333–366. (link)
[2] Hamley, I.W., Angew. Chem. Int. Ed. 2007, 46, 8128–8147. (link)
[3] Adler, J.; Huster, D., Phys. Chem. Chem. Phys. 2017, 19, 1839–1846. (link)
[4] Funtan, S.; Evgrafova, Z.; Adler, J.; Huster, D.; Binder, W.H., Polymers 2016, 8, 178. (link)