Copper(II) complexation by pituitary adenylate cyclase activating polypeptide fragments.

Results are reported from potentiometric and spectroscopic (UV-Vis, CD, and ESR) studies of the protonation constants and Cu2+ complex stability constants of pituitary adenylate cyclase activating polypeptide fragments (HSDGI-NH2, TDSYS-NH2, RKQMAVKKYLAAVL-NH2). With HSDGI-NH2, the formation of a dimeric complex Cu2H-2L2 was found in the pH range 5-8, in which the coordination of copper(II) is glycylglycine-like, while the fourth coordination site is occupied by the imidazole N3 nitrogen atom, forming a bridge between two copper(II) ions. The formation of dimeric species does not prevent the deprotonation and coordination of the amide nitrogen, and in pH above 8 the CuH-2L complex is formed. Aspartic acid in the third position of peptide sequence stabilizes the CuH-2L species and prevents the coordination of a fourth nitrogen donor. Aspartic acid residue in the second position of TDSYS-NH2 stabilizes the CuL (2N) complex but does not prevent deprotonation and binding of the second and third peptide nitrogens to give 3N and 4N complexes at higher pH. The tetradecapeptide amide forms with copper(II) ions unusually stable 3N and 4N complexes compared to pentaalanine amide.     

Ab initio simulations of Cu binding sites on the N-terminal region of prion protein

The human prion protein binds Cu²⁺ ions in the octarepeat domain of the N-terminal tail up to full occupancy at pH 7.4. Recent experiments have shown that the HGGG octarepeat subdomain is responsible for holding the metal bound in a square-planar configuration. By using first principle ab initio molecular dynamics simulations of the Car–Parrinello type, the coordination of copper to the binding sites of the prion protein octarepeat region is investigated. Simulations are carried out for a number of structured binding sites. Results for the complexes Cu(HGGGW)(wat), Cu(HGGG), and [Cu(HGGG)]₂ are presented. While the presence of a Trp residue and a water molecule does not seem to affect the nature of the copper coordination, high stability of the bond between copper and the amide nitrogen of deprotonated Gly residues is confirmed in all cases. For the more interesting [Cu(HGGG)]₂ complex, a dynamically entangled arrangement of the two domains with exchange of amide nitrogen bonds between the two copper centers emerges, which is consistent with the short Cu–Cu distance observed in experiments at full copper occupancy.     

Coordination abilities of neurokinin A and its derivative and products of metal-catalyzed oxidation

The classical tachykinins, substance P, neurokinin A and neurokinin B are predominantly found in the nervous system where they act as neurotransmitters and neuromodulators. Significantly reduced levels of these peptides were observed in neurodegenerative diseases and it may be suggested that this reduction may also result from the copper(II)-catalyzed oxidation. The studies of the interaction of copper(II) with neurokinin A and the copper(II)-catalyzed oxidation were performed. Copper(II) complexes of the neurokinin A (His-Lys-Thr-Asp-Ser-Phe-Val-Gly-Leu-Met-NH₂) and acetyl-neurokinin A (Ac-His-Lys-Thr-Asp-Ser-Phe-Val-Gly-Leu-Met-NH₂) were studied by potentiometric, UV-Vis (UV-visible), CD (circular dichroism) and EPR spectroscopic methods to determine the stoichiometry, stability constants and coordination modes in the complexes formed. The histidine residue in first position of the peptide chain of neurokinin A coordinates strongly to Cu(II) ion with histamine-like {NH₂, N_Im} coordination mode. With increasing of pH, the formation of a dimeric complex Cu₂H₂L₂ was found but this dimeric species does not prevent the deprotonation and coordination of the amide nitrogens. In the Ac-neurokinin A case copper(II) coordination starts from the imidazole nitrogen of the His; afterwards three deprotonated amide nitrogens are progressively involved in copper coordination. To elucidate the products of the copper(II)-catalyzed oxidation of the neurokinin A and Ac-neurokinin A, liquid chromatography-mass spectrometry (LC-MS) method and Cu(II)/hydrogen peroxide as a model oxidizing system were employed.Oxidation target for both studied peptides is the histidine residue coordinated to the metal ions. Both peptides contain Met and His residues and are very susceptible on the copper(II)-catalyzed oxidation.     

A multinuclear nmr relaxation study of the interaction of divalent metal ions with l-aspartic acid

Carbon-13 spin-lattice relaxation times, T₁, have been measured for aqueous solutions of L-aspartic acid, L-alanine, O-phospho-L-serine, and 2-mercapto-L-succinic acid in the presence of the paramagnetic metal ions, Cu²⁺ and Mn²⁺, and Mg²⁺ as a diamagnetic control, at ambient temperature and neutral pH. Nitrogen-15, oxygen-17 and proton relaxation times were also obtained for L-aspartic acid and phosphorus-31 relaxation times for O-phospho-L-serine under similar conditions. The structures of these complexes in solution were determined from the various metal ion-nuclei distances calculated from the paramagaetically-induced relaxation. These results indicate that the Cu²⁺ interaction with L-aspartic acid is through α-amino and β-carboxyl groups while Mn²⁺ coordinates most strongly through α-and β-carboxyl groups, with the possibility of a weak interaction through the amino group.An examination of the coordination of these divalent metal ions to an analog of L-aspartic acid in which the β-carboxyl group is replaced by a phosphate group (O-phospho-L-serine) indicated that Cu²⁺ coordination is now probably through the α-amino and phosphate groups, while this analog is a monodentate ligand for Mn²⁺ coordinating through the phosphate group. Removal of the β-carboxyl group (L-alanine) also results in Cu²⁺ coordination through the α-carboxyl and α-amino groups, and the same ligand interactions are observed with Mn²⁺. Replacement of the α-amino group of L-aspartic acid with an - SH group (2-mercapto-L-succinate) is sufficient to eliminate any specific coordination with either Cu²⁺ or Mn²⁺.     

Copper(II), nickel(II) and zinc(II) complexes of N-acetyl-His-Pro-His-His-NH2: equilibria, solution structure and enzyme mimicking

The copper(II), nickel(II) and zinc(II) binding ability of the multi-histidine peptide N-acetyl-His-Pro-His-His-NH₂ has been studied by combined pH-potentiometry and visible, CD and EPR spectroscopies. The internal proline residue, preventing the metal ion induced successive amide deprotonations, resulted in the shift of this process toward higher pH values as compared to other peptides. The metal ions in the parent [ML]²⁺ complexes are exclusively bound by the three imidazole side chains. In [CuH₋₁L]⁺, formed between pH 6-8, the side chains of the two adjacent histidines and the peptide nitrogen between them are involved in metal ion binding. The next deprotonation results in the proton loss of the coordinated water molecule (CuH₋₁L(OH)). The latter two species exert polyfunctional catalytic activity, since they possess superoxide dismutase-, catecholase- (the oxidation of 3,5-di-tert-butylcatechol) and phosphatase-like (transesterification of the activated phosphoester 2-hydroxypropyl-4-nitrophenyl phosphate) properties. On further increase of the pH rearrangement of the coordination sphere takes place leading to the [CuH₋₃L]⁻ species, the deprotonated amide nitrogen displaces a coordinated imidazole nitrogen from the equatorial position of the metal ion. The shapes of the visible and CD spectra reflect a distorted arrangement of the donor atoms around the metal ion. In presence of zinc(II) the species [ZnL]²⁺ forms only above pH 6, which is shortly followed by precipitation. On the other hand, the [NiL]²⁺ complex is stable over a wide pH range, its deprotonation takes place only above pH 8. At pH 10 an octahedral NiH₋₂L species is present at first, which transforms slowly to a yellow square planar complex.     

Interaction of copper(II) and nickel(II) with a bis-amide ligand functionalized with pyridine moieties: Thermodynamic and spectroscopic studies in aqueous solution

We synthesized a new bis-amide ligand derived from the l(+)-tartaric acid. We then determined its protonation constants and the stability constants of the copper(II) and nickel(II) chelates by potentiometry as well as ESI-MS and UV-Vis spectroscopy. We found that both metal ions are able to induce the deprotonation and the coordination of an amide nitrogen donor atom. In the case of copper complexes, the data show the formation of two major species: Cu₂(L₂H₋₃)⁺ and Cu₂(LH₋₄). EPR and XAS experiments led us to precise the relative structure of these compounds. In Cu₂(L₂H₋₃)⁺, each metal center is coordinated by pyridinic and amidic nitrogen atoms of one ligand and by nitrogen and oxygen atoms from pyridine and hydroxyl moieties from the other one. In Cu₂(LH₋₄), the copper centers are coordinated by pyridinic and amidic nitrogen atoms, as well as a deprotonated hydroxyl group of the ligand. In this latter complex, the lower value of the Cu-Cu distance determined from EXAFS experiments and compared to the one of the solid species likely involve the formation of an exogeneous hydroxyl bridge between the two copper centers. With Ni(II) ions, the only one major species is the mononuclear Ni(LH₋₂) complex, in which Ni(II) is held in an octahedral environment with the metal center chelated by the two pyridinic and the two amidic nitrogen atoms, and two oxygen atoms from water molecules.     

Mononuclear copper(II) complexes of alloferons 1 and 2: a combined potentiometric and spectroscopic studies

Mononuclear copper(II) complexes of the alloferon 1 His-Gly-Val-Ser-Gly-His-Gly-Gln-His-Gly-Val-His-Gly, alloferon 2 Gly-Val-Ser-Gly-His-Gly-Gln-His-Gly-Val-His-Gly, Ac-alloferon 1 Ac-His-Gly-Val-Ser-Gly-His-Gly-Gln-His-Gly-Val-His-Gly and Ac-alloferon 2 Ac-Gly-Val-Ser-Gly-His-Gly-Gln-His-Gly-Val-His-Gly have been studied by potentiometric, UV-vis, CD and EPR spectroscopic methods. The potentiometric and spectroscopic data shows that acetylation of the amino terminal group induces significant changes in the coordination properties of the Ac-alloferons 1 and 2 compared to the alloferons 1 and 2, respectively. The presence of four (Ac-alloferon 1) or three (Ac-alloferon 2) 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 coordination of amide functions and the CuH₋₃L species with {N_Im, 3N⁻} bonding mode at pH above 8 are formed. The N-terminal amino group of the alloferons 1 and 2 takes part in the coordination of the metal ion and the 4N complex with {NH₂, 3N_Im} coordination mode dominates at physiological pH 7.4 for alloferon 1 and the 3N {NH₂, CO, 2N_Im} binding mode for alloferon 2. However, at higher pH values sequential amide nitrogens are deprotonated and coordinated to copper(II) ions.     

Modeling of copper(II) sites in proteins based on histidyl and glycyl residues

The complexes between copper(II) and four synthetic tetrapeptides bearing a single histidine residue within the sequence (AcHGGG, AcGHGG, AcGGHG and AcGGGH, respectively), have been investigated by potentiometric and spectroscopic methods (UV-Vis, circular dichroism and electron paramagnetic resonance). Potentiometric studies in the pH range 4-12 allowed identification and quantitative determination of the species present in solution for each copper-peptide complex. In all cases, upon raising pH, copper(II) coordination starts from the imidazole nitrogen of the His; afterwards three deprotonated amide nitrogens are progressively involved in copper coordination, except in the case of AcGHGG. Based on the potentiometric and spectroscopic results, detailed molecular structures are proposed for the dominant copper(II) tetrapeptide species existing in solution, either at neutral or alkaline pH. The structural consequences of the presence and of the location of a unique histidine residue within the tetrameric sequence are specifically analyzed. Results are discussed in relation to the modeling of copper(II) binding sites in proteins, particular emphasis being devoted to the copper complexes of the prion protein.     

Structure and stability of the CuII complexes with tandem repeats of the chicken prion

Prion protein (PrP) misfolding is one of the pivotal issues in understanding the rudiments of neurodegenerative disorders. The conformational change of mammalian cellular PrP to scrapie PrP is caused by an unknown agent, but there is reasonable evidence supporting the key role of copper ions in this process. The structure of the avian PrP was found to be very similar to the mammalian protein, although there is only 30% homology in the secondary structure. This work shows that copper ions are very effectively bound by hexarepeat fragments of chicken prion protein, although not as effectively as it was found in the case of mammalian protein. By means of potentiometric and spectroscopic techniques (nuclear magnetic resonance, circular dichroism, UV-vis, and electronic paramagnetic resonance), it was shown that Cu(II) ions coordinate to the chicken PrP hexapeptide domain in physiological pH via imidazole nitrogen donors of His residue(s). The binding pattern changes the structure of peptide involved, indicating a possible impact of Cu(II) ions in the biology and pathology of nonmammalian PrP, which could be similar to that found for mammalian PrP. The present study shows that, similar to the human prion octapeptide repeats, chicken prion hexapeptide repeats might bind copper ions in two different ways, depending on the number of repeats and metal/ligand molar ratio: (i) an intra-repeat coordination mode in which copper ion is chelated by His imidazole and deprotonated amide nitrogen in monomeric peptide and (ii) an inter-repeat coordination mode in which a polymeric peptide ligand (dimer and trimer) forms polyimidazole complexes that are very stable at physiological pH. Two proline residues inserted into the hexapeptide unit have a critical impact on the metal-binding ability.     

Impact of ring size on the copper(II) coordination abilities of cyclic tetrapeptides

A new, 14-membered, tetraza cyclic tetrapeptide containing histidine and lysine side-chains, c(β³homoLysdHisβ-AlaHis), was designed, synthesized and characterized; its copper(II) binding properties were investigated in dependence of pH by potentiometric and spectroscopic methods. In line with previous studies of similar systems, the progressive involvement of amide nitrogens in copper(II) coordination was evidenced for pH values greater than 6. At physiological pH the dominant species consists of a copper(II) center coordinated by two amide nitrogens, an imidazole nitrogen and a water molecule. In contrast, at pH values higher than 8.7, a copper(II) coordination environment consisting of four amide nitrogens in the equatorial plane and the axial imidazole ligands is formed as clearly indicated by spectroscopic data and theoretical calculations. The behavior of this 14-membered cyclic tetrapeptide is compared to that of its 12-membered cyclic analog, particular attention being paid to the effects of ring size on the respective copper(II) binding abilities.     

Coordination abilities of substituted β-aminophosphonates towards Cu and Zn ions

The formation constants of equimolar and bis-chelate copper(II) and zinc(II) complexes with three aliphatic and four aromatic-substituted β-aminophosphonates have been determined in water solution by potentiometric studies. Spectroscopic parameters clearly indicate involvement of {NH₃, PO₃ ²⁻} in both metal ions coordination. The comparison of the stability constants reveals slightly higher coordination power of the aliphatic-substituted β-aminophosphonic acids, which may be due to the higher basicity of their amino groups. All studied ligands are more effective in Cu²⁺ and Zn²⁺ coordination than phosphonic analogue of simple β-amino acid.     

Copper(II) complexes of Neobelliera Bullata Trypsin Modulating Oostatic Factor and its analogues

The stoichiometry, stability constants and solution structure of the complexes formed in the reaction of copper(II) with hexapeptide NPTNLH, i.e. the Neobelliera Bullata Trypsin Modulating Oostatic Factor (Neb-TMOF), and its analogues DPTNLH, Ac-NPTNLH and Ac-DPTNLH have been determined by potentiometric, UV-visible, CD and EPR spectroscopic methods. Upon raising pH for Ac-NPTNLH and Ac-DPTNLH peptides, copper(II) coordination starts from the imidazole nitrogen of the His⁶; afterwards three deprotonated amide nitrogens are progressively involved in metal ions coordination. In a wide pH range of 4.5-8.5 for the NPTNLH and DPTNLH ligands the CuL complex dominates with the imidazole nitrogen of His⁶ coordinated to form a macrochelate. The N-terminal amino group of the NPTNLH and DPTNLH peptides takes part in the coordination of the metal ion in the CuL, CuH₋₁L and CuH₋₂L complexes. However, at pH above 9 the CuH₋₃L complex with the {N_Im, 3N⁻} coordination mode is formed. For the CuH₋₂L complex the spectroscopic data clearly indicate the 4N {NH₂, CO or COO⁻, 2N⁻, N_Im} bonding mode with the axial coordination of the N-terminal amine group to the metal ion.     

Binding of metal ions to polysaccharides. VI. Spectroscopic and potentiometric studies of the binding of copper(II) and nickel(II) to some monosaccharide units in acidic,neutral and alkaline aqueous media

Binding of Cu²⁺ and Ni²⁺ to glucosamine, N-acetyl- glucosamine and other derivatives of glucose was investigated in acidic, neutral and alkaline aqueous media using H⁺ and Cu²⁺ potentiometry and ligand- field and ESR spectroscopy. In neutral medium, site binding with copper(II) and nickel(II) occurs when the monosaccharide possesses a potentially coordinating amine or charged group not attached to C-1. At high pH, a coordination entity is only formed if the C-1 hydroxyl group can be deprotonated and other stabilizing groups are present. The role of groups attached to C-1 reflects the different behaviour of monosaccharides compared with polysaccharides.     

Model studies on the coordination of copper in biological systems. The deprotonated peptide nitrogen as a potential binding site for copper(II)

1. A large number of potentially bidentate and tridentate amides, X-Y-CONH-Z, were used as model ligands to investigate the complex formation of Cu(II) with the deprotonated peptide nitrogen in biological molecules. A combination of potentiometric titration, spectrophotometry and electron paramagnetic resonance was applied to analyse the structure of the Cu(II) chelates formed at neurtal and basic pH. 2. By systematic variation of the primary binding function X, the ring size of the chelate, and the spatial properties of the C-terminal and N-terminal substituents, three classes of amide ligands could be established with reference to their capacity for Cu(II)-induced deprotonation of NHCO and metal binding. 3. Under physiological conditions of pH, peptide (class A) chelates are only formed by those bidentate amide ligands with X being an imidazole (sp2) nitrogen or a terminal (sp3) amino nitrogen. Mercaptide sulfur must also be considered to belong in this group of strong copper(II)-binding sites, but in our low-molecular-weight model ligands the redox equilibrium 2 Cu(II) + 2 RSH in equilibrium or formed from 2 CU(II) + RSSR + 2 H+ interferes, yielding insoluble Cu(I)-S polymers above pH 4. In addition to the unidentate binding strength of X, entropy effects play an important role. Depending on whether X is an imidazole or amino nitrogen, only five-membered or six-membered monocyclic chelate structures respectively cause coordination of the deprotonated peptide function. 4. Biuret (class B) Cu(II) chelates are only formed under non-physiological conditions at pH > 11.5 producing the well known violet chromophores CuIIN4(-). In general these complexes, which also include the Cu(II) biguanides, show a clearly resolved electron paramagnetic resonance spectrum with nitrogen superhyperfine structure. 5. A third class of peptide model ligands (class C) consists of those amides where the CuII-X bond does not provide enough thermodynamic stability. The binding site of these class C amides includes functional groups such as carboxylate (COO-), methionine sulfur (RSR'), aliphatic or aromatic hydroxyl (OH) and amide nitrogen (NHCO) itself. When X is a pyridine (sp2) nitrogen or an amino (sp3) nitrogen, NHCO deprotonation is only promoted in five-membered but not six-membered ring chelates. On the other hand, a combination of COO- and NH2, as in asparagine, will allow deprotonation of NHCO in the presence of Cu(II). And third, despite a pronounced unidentate affinity of the imidazole nitrogen for Cu(II), N-acetylhistamine acts as a class C amine, in contrast to imidazolylacetamide, which forms a stable Cu(II) peptide chelate. This difference in Cu binding is explained on the basis of space-filling models. These clearly demonstrate that in the case of the 2:1 complex of Cu(II) with N-acetylhistamine, the planarity of the ionised peptide function can not be retained in a square planar arrangement of the two amide ligands around the copper center.     

Copper(II) complexes with peptide fragments encompassing the sequence 122-130 of human doppel protein

Copper(II) complexes of the peptide fragment (Dpl122-130) encompassing the sequence 122-130 of human doppel protein were characterized by potentiometric, UV-Visible, CD and EPR spectroscopic methods. An analogous peptide, in which the aspartate residue was substituted by an asparagine amino acid, was synthesized in order to provide evidence on the possible role of carboxylate group in copper(II) coordination. It was found that the carboxylic group is directly involved in copper(II) coordination at acidic pH, forming the CuLH₂ species with Dpl122-130. This copper(II) complex displayed EPR parameters very similar to those of the analogous complex with the whole doppel protein. At pH higher than 7, the complexes showed magnetic parameters similar to those of the major species of protein formed in the pH range 7-8, with the metal coordination environment consisting of one imidazole and three amide nitrogen atoms. The comparison of Cu-Dpl122-130 binding constant values with those of the prion peptide fragments (PrP106-114), showed that doppel peptide had a higher metal binding affinity at acidic pH whereas the prion peptide fragment binds the metal tightly at physiological pH.     

The configuration of the Cu2+ binding region in full-length human prion protein

The cellular prion protein (PrP^C) is a Cu²⁺ binding protein connected to the outer cell membrane. The molecular features of the Cu²⁺ binding sites have been investigated and characterized by spectroscopic experiments on PrP^C-derived peptides and the recombinant human full-length PrP^C (hPrP-[23-231]). The hPrP-[23-231] was loaded with ⁶³Cu under slightly acidic (pH 6.0) or neutral conditions. The PrP^C/Cu²⁺-complexes were investigated by extended X-ray absorption fine structure (EXAFS), electron paramagnetic resonance (EPR), and electron nuclear double resonance (ENDOR). For comparison, peptides from the copper-binding octarepeat domain were investigated in different environments. Molecular mechanics computations were used to select sterically possible peptide/Cu²⁺ structures. The simulated EPR, ENDOR, and EXAFS spectra of these structures were compared with our experimental data. For a stoichiometry of two octarepeats per copper the resulting model has a square planar four nitrogen Cu²⁺ coordination. Two nitrogens belong to imidazole rings of histidine residues. Further ligands are two deprotonated backbone amide nitrogens of the adjacent glycine residues and an axial oxygen of a water molecule. Our complex model differs significantly from those previously obtained for shorter peptides. Sequence context, buffer conditions and stoichiometry of copper show marked influence on the configuration of copper binding to PrP^C. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Computational studies of Cu(II)[peptide] binding motifs: Cu[HGGG] and Cu[HG] as models for Cu(II) binding to the prion protein octarepeat region

The binding of Cu(II) to the prion protein is investigated by computations at the B3LYP level of theory on models of the octarepeat domain of the prion protein. The models incorporate the functionality of the glycine (G) and histidine (H) residues which occur in the octarepeat domain, PHGGGWGQ. The copper complexes are designated Cu[HG] and Cu[HGGG]. Coordination to the metal via the imidazole ring of the histidine, the amide carbonyl groups, and the backbone nitrogen atom of the amide groups were examined, as well as several protonation/deprotonation states of each structure. EPR and CD titration experiments suggest that the octarepeat segments of the unstructured N-terminal domain of prion protein can bind Cu(II) in a 1:1 Cu-to-octarepeat ratio. The results identify the extent to which the Cu(II) facilitates peptide backbone deprotonation, and the propensity of binding in the forward (toward the C-terminus) direction from the anchoring histidine residue. A plausible mechanism is suggested for changing from amide O-atom to deprotonated amide N-atom coordination, and for assembly of the observed species in solutions of Cu[PrP] and truncated models of it. A structure is proposed which has the N2O2 coordination pattern for the minor component observed experimentally by EPR spectroscopy for the Cu[HGGG] model. The most stable neutral Cu[HGGG] structure found, with coordination environment N3O1, corresponds to that observed for Cu[HGGGW] and Cu[HGGG] both in the solid state and as the major component in solution at neutral pH.     

Cu(II) interaction with N-terminal fragments of human and mouse beta-amyloid peptide

The stoichiometry, stability constants and solution structure of the complexes formed in the reaction of copper(II) with N-terminal fragments of human and mouse beta-amyloid peptide, 1-6, 1-9, 1-10 have been determined by potentiometric, UV/VIS, CD and EPR spectroscopic methods. The fragments 1-9 and 1-10 form complexes with the same coordination modes as the fragments 1-6. The coordination of the metal ion for human and mouse fragments starts from the N-terminal Asp residue which stabilizes significantly the 1N complex as a result of chelation through the beta-carboxylate group. In a wide pH range of 4-10, the imidazole nitrogen of His(6) is coordinated to form a macrochelate. Results show that, in the pH range 5-9 the human fragments form the complex with different coordination mode compared to that of the mouse fragments. The low pK(1)(amide) values (approximately 5) obtained for the mouse fragments may suggest the coordination of the amide nitrogen of His(6) while in case of the human fragments the coordination of the amide nitrogen of Ala(2) is suggested. The replacement of glycine by the arginine residue in the fifth position of the beta-amyloid peptide sequence changes the coordination modes of a peptide to metal ion in the physiological pH range. In a wide pH (including physiological) range the mouse fragments of beta-amyloid peptide are much more effective in Cu(II) binding than the human fragments.     

Metal binding ability of hypermodified nucleosides of t-RNA. Potentiometric and spectroscopic studies on the metal complexes of N-[(9-β-D-ribofuranosylpurin-6-yl)carbamoyl] threonine

Copper(II), nickel(II), zinc(II), manganese(II), and magnesium(II) complexes of t⁶A (N-[9-β-D-ribofuranosylpurin-6-yl)carbamoyl] threonine and t⁶Ade (N⁶(threoninocarbonyl)adenine) were studied by potentiometric and spectroscopic methods. It was found that t⁶Ade has three dissociable protons in the accessible pH range (N¹ and N⁹ of purine and carboxylate), while only two pK values are characteristic of t⁶A. Magnesium(II) and manganese(II) do not interact effectively with these ligands, but copper(II) and nickel(II) ions form very stable complexes with the coordination of purine N¹, deprotonated amide nitrogen, and carboxy late oxygen donors.     

Coordination abilities of the 1-16 and 1-28 fragments of beta-amyloid peptide towards copper(II) ions: a combined potentiometric and spectroscopic study

Stoichiometry, stability constants and solution structures of the copper(II) complexes of the (1-16H), (1-28H), (1-16M), (1-28M), (Ac-1-16H) and (Ac-1-16M) fragments of human (H) and mouse (M) beta-amyloid peptide were determined in aqueous solution in the pH range 2.5-10.5. The potentiometric and spectroscopic data (UV-Vis, CD, EPR) show that acetylation of the amino terminal group induces significant changes in the coordination properties of the (Ac-1-16H) and (Ac-1-16M) peptides compared to the (1-16H) and (1-16M) fragments, respectively. The (Ac-1-16H) peptide forms the 3N [N(Im)(6), N(Im)(13), N(Im)(14)] complex in a wide pH range (5-8), while for the (Ac-1-16M) fragment the 2N [N(Im)(6), N(Im)(14)] complex in the pH range 5-7 is suggested. At higher pH values sequential amide nitrogens are deprotonated and coordinated to copper(II) ions. The N-terminal amino group of the (1-16) and (1-28) fragments of human and mouse beta-amyloid peptide takes part in the coordination of the metal ion, although, at pH above 9 the complexes with the 4N [N(Im), 3N(-)] coordination mode are formed. The phenolate -OH group of the Tyr(10) residue of the human fragments does not coordinate to the metal ion.