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Copper(II) complexes of the pentapeptides Ac-HisAlaHisValHis-NH2, Ac-HisValHisAlaHis-NH2, Ac-HisProHisAlaHis-NH2, Ac-HisAlaHisProHis-NH2, Ac-HisGlyHisValHis-NH2 and Ac-HisValHisGlyHis-NH2 have been studied by potentiometric, UV-Vis, CD and EPR spectroscopic methods. It has been found that the pentapeptides are efficient ligands for the complexation with copper(II) and exhibit an outstanding versatility in the co-ordination geometry of complexes. The presence of three histidyl residues provides a high possibility for the formation of macrochelates via the exclusive binding of imidazole-N donor atoms. The macrochelation suppresses, but cannot preclude the deprotonation and metal ion co-ordination of amide functions and the species [CuH(-2)L] and [Cu2H(-4)L] predominate at physiological pH in equimolar solutions and in the presence of excess metal ions, respectively. It is also clear from the data that both C-terminal and internal histidyl residues can work as the anchoring sites for metal binding and subsequent amide deprotonation resulting in the formation of co-ordination isomers and dinuclear species in equimolar solutions and in the presence of excess metal ions, respectively. In more alkaline solutions (pH approximately 10) a third amide function can be deprotonated and co-ordinated in the species [CuH(-3)L]- with (N-,N-,N-,N(im)) co-ordination. The dinuclear species [Cu2H(-5)L]- and [Cu2H(-6)L](2-) containing hydroxide ions and/or imidazolato bridges are formed at high pH in the presence of excess of metal ions. The insertion of one proline into the sequence preceding histidyl residues hinders the deprotonation of amide functions at that site and the formation of only mononuclear complexes was observed with these peptides.

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Chromatin proteins are believed to represent reactive sites for metal ion binding. We have synthesized the 31 amino acid peptide Ac-NSFVNDIFERIAGEASRLAHYNKRSTITSRE-NH2, corresponding to the 63-93 fragment of the histone H2B and studied its interaction with Cu(II) and Ni(II). Potentiometric and spectroscopic studies (UV-vis, CD, NMR and EPR) showed that histidine 21 acts as an anchoring binding site for the metal ion. Complexation of the studied peptide with Cu(II) starts at pH 4 with the formation of the monodentate species CuH2L. At physiological pH values, the 3N complex (N(Im), 2N(-)), CuL is favoured while at basic pH values the 4N (N(Im), 3N(-)) coordination mode is preferred. Ni(II) forms several complexes with the peptide starting from the distorted octahedral NiH2L at about neutral pH, to a square planar complex where the peptide is bound through a (N(Im), 3N(-)) mode in an equatorial plane at basic pH values. These results could be important in revealing more information about the mechanism of metal induced toxicity and carcinogenesis.

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The N- and C-terminal blocked hexapeptide Ac-Leu-Ala-His-Tyr-Asn-Lys-amide (LAHYNK) representing the 80–85 fragment of histone H2B was synthesized and its interactions with Cu(II) and Ni(II) ions were studied by potentiometric, UV-Vis, CD, EPR, and NMR spectroscopic techniques in solution. Our data reveal that the imidazole N(3) nitrogen atom is the primary ligating group for both metal ions. Sequential amide groups deprotonation and subsequent coordination to metal ions indicated an {Nimidazole, 3Namide} coordination mode above pH∼9, in all cases. In the case of Cu(II)-peptide system, the almost exclusive formation of the predominant species CuL in neutral media accounting for almost 98% of the total metal ion concentration at pH 7.3 strongly indicates that at physiological pH values the sequence -LAHYNK- of histone H2B provides very efficient binding sites for metal ions. The imidazole pyrrole N(1) ionization (but not coordination) was also detected in species CuH−4L present in solution above pH ∼ 11.

In order to reveal more information about the toxicity caused by metals and furthermore their influence to the physiological metabolism of the cell, the hexapeptide model Ac-ThrTyrThrGluHisAla-am representing the C-terminal 71-76 fragment of histone H4 which lies into the nucleosome core, was synthesized. A combined pH-metric and spectroscopic UV-VIS, EPR, CD and NMR study of Ni(II) and Cu(II) binding to the blocked hexapeptide, revealed the formation of octahedral complexes involving imidazole nitrogen of histidine, at pH 5 and pH 7 for Cu(II) and Ni(II) ions respectively. In basic solutions a major square-planar 4 N Ni(II)-complex, adopting a {N(Im), 3N(-)} coordination mode, was formed. In the case of Cu(II) ions, a 3 N complex, involving the imidazole nitrogen of histidine and two deprotonated amide nitrogens of the backbone of the peptide, at pH 7 and a series of 4 N complexes starting at pH 6.5, were suggested. In addition Ni(II)-mediated hydrolysis of the peptide bond-Tyr-Thr was evident following our experimental data.