Copper(II) complexes of the Neb-colloostatin analogues containing histidine residue structure stability biological activity

Abstract

Neb-colloostatin was discovered during the isolation of Neb-TMOF from the ovary of the grey fleshfly Neobellieria bullata. Its amino acid sequence was determined as SIVP4LGLP8VP10IGP13IVVGP18R. The analogues with point mutation P4H and P8H were synthesized and their copper(II) complexes were studied by potentiometry, UV–Vis, circular dichroism (CD), and electron paramagnetic resonance (EPR) spectroscopic and mass spectrometry (MS) methods. To obtain a complete complex speciation 1:1 and 2:1 metal-to-ligand molar ratios for both peptides were studied. At physiological pH (7.4), both peptides form mononuclear the CuH-1L complex with 3N{NH2, N−, NIm} coordination mode. For the P4H and P8H peptides at high pH (∼11) the binding sites of copper(II) ions are quite different. For the CuH-4L complex of the P4H peptide the 4N{4N−} coordination mode is dominante, while for the P8H peptide the 4N{NIm, 3N−} binding site towards N-termini is formed with (6,5,5) chelate ring. At high pH the P8H peptide cannot form the 4N {NH2, 3N−} complex because of the presence of the Pro4 residue in the amino acid sequence (a break point in the metal coordination). At pH 7.4 dinuclear the Cu2H-4L complexes dominate with suggested the 3N{NH2, 2N−}3N{NIm, 2N−}binding sites where the 3N{NIm, 2N−} coordination mode for the P4H peptide towards C-termini, and for P8H peptide towards N-termini are formed. The induction of apoptosis in vivo in Tenebrio molitor cells by the ligands and their copper(II) complexes at pH 7.4 was studied. The systems studied have lower apoptotic properties compared to those of Neb-colloostatin and its copper(II) complex at pH 7.4.