Copper(II) Coordination Abilities of the Tau Protein's N-Terminus Peptide Fragments: A Combined Potentiometric, Spectroscopic and Mass Spectrometric Study.

Márton Lukács,Giuseppe Pappalardo,Ágnes Grenács,Giuseppe Di Natale,Katalin Várnagy,Norbert Lihi,Imre Sóvágó,Györgyi Szunyog,Tiziana Campagna,Valeria Lanza,Csilla Kállay,Giovanni Tabbi

doi:10.1002/cplu.201900504

Abstract

Copper(II) complexes of the N-terminal peptide fragments of tau protein have been studied by potentiometric and various spectroscopic techniques (UV-vis, CD, ESR and ESI-MS). The octapeptide Tau(9-16) (Ac-EVMEDHAG-NH2 ) contains the H14 residue of the native protein, while Tau(26-33) (Ac-QGGYTMHQ-NH2 ) and its mutants Tau(Q26K-Q33K) (Ac-KGGYTMHK-NH2 ) and Tau(Q26K-Y29A-Q33K) (Ac-KGGATMHK-NH2 ) include the H32 residue. To compare the binding ability of H14 and H32 in a single molecule the decapeptide Ac-EDHAGTMHQD-NH2 (Tau(12-16)(30-34)) has also been synthesized and studied. The histidyl residue is the primary metal binding site for metal ions in all the peptide models studied. In the case of Tau(9-16) the side chain carboxylate functions enhance the stability of the M-Nim coordinated complexes compared to Tau(26-33) (logK(Cu-Nim )=5.04 and 3.78, respectively). Deprotonation and metal ion coordination of amide groups occur around the physiological pH range for copper(II). The formation of the imidazole- and amide-coordinated species changes the metal ion preference and the complexes formed with the peptides containing the H32 residue predominate over those of H14 at physiological pH values (90 %-10 %) and in alkaline samples (96 %-4 %).

Highlights

Copper(II) complexes of the N-terminal peptide fragments of primary metal binding site for metal ions in all the peptide tau protein have been studied by potentiometric and various models studied
A comprehensive mass spectrometry screening on cerebrospinal fluid samples of Protonation constants of the peptides have been determined by potentiometric measurements and the data are listed in patients with Alzheimer disease revealed an increased level of Table 1
Dissociation of the His32ÀGln33 amide bond is favoured over dissociation of the His14ÀAla15 bond suggesting the copper(II) preference for binding to the His32 binding site

Summary

Results and Discussion

In almost all previous complexation studies the peptide fragments of the microtubule-binding domain were investi- Protonation Equilibria of the Peptides gated, histidines are present in the N-terminal. 44 works and is indicative of conformational changes mainly related to the copper(II) coordination by imidazole side chain of histidine residue and the amide nitrogens.[27] Interestingly, the 47 different far-UV CD spectra of the Cu(II)ÀTau(26-33) system reveal the appearance of this distinctive CD band profile at 49 lower pH (pH 5–6) than those reported in the Cu(II)ÀTau(9-16) system thereby suggesting an enhanced stability of the (Nim,NÀ,NÀ) coordination mode in the former metal-peptide system. Suggesting the preference for copper(II) binding at His sites at 1 : 1 metal to peptide ratio and basic pH values All these results are in a good agreement with those the major species between pH 5–7 and its stoichiometry obtained for the small one-histidine fragments as it was strongly supports the involvement of both imidazole moieties demonstrated by Figure 4. The MS/MS spectra recorded for the -Cu (II)ÀTau(12-16)(30-34) system at 1 : 1 metal to peptide ratio (Figure 7) indicate that the peak corresponding to the Cu-b9 fragment was the principal fragment observed when the

Conclusion

13 Peptide Synthesis

A Bruker Elexsys E500 CW-ESR spectrometer driven by a PC running