The extent to which the replacement of a ribose unit by a 2'-deoxyribose unit influences the acid-base properties of nucleotides has not hitherto been determined in detail. In this study, by potentiometric pH titrations in aqueous solution, we have measured the acidity constants of the 5'-di- and 5'-triphosphates of 2'-deoxyguanosine [i.e., of H(2)(dGDP)(-) and H(2)(dGTP)(2-)] as well as of the 5'-mono-, 5'-di-, and 5'-triphosphates of 2'-deoxyadenosine [i.e., of H(2)(dAMP)(+/-), H(2)(dADP)(-), and H(2)(dATP)(2-)]. These 12 acidity constants (of the 56 that are listed) are compared with those of the corresponding ribose derivatives (published data) measured under the same experimental conditions. The results show that all protonation sites in the 2'-deoxynucleotides are more basic than those in their ribose counterparts. The influence of the 2'-OH group is dependent on the number of 5'-phosphate groups as well as on the nature of the purine nucleobase. The basicity of N7 in guanine nucleotides is most significantly enhanced (by about 0.2 pK units), while the effect on the phosphate groups and the N1H or N1H(+) sites is less pronounced but clearly present. In addition, (1)H NMR chemical shift change studies in dependence on pD in D(2)O have been carried out for the dAMP, dADP, and dATP systems, which confirmed the results from the potentiometric pH titrations and showed the nucleotides to be in their anti conformations. Overall, our results are not only of relevance for metal ion binding to nucleotides or nucleic acids, but also constitute an exact basis for the calculation, determination, and understanding of perturbed pK(a) values in DNAzymes and ribozymes, as needed for the delineation of acid-base mechanisms in catalysis.

The interaction between metal ions and nucleotides is well characterized, as is their importance for metabolic processes, e.g. in the synthesis of nucleic acids. Hence, it is surprising to find that no detailed comparison is available of the metal ion-binding properties between nucleoside 5'-phosphates and 2'-deoxynucleoside 5'-phosphates. Therefore, we have measured here by potentiometric pH titrations the stabilities of several metal ion complexes formed with 2'-deoxyadenosine 5'-monophosphate (dAMP2-), 2'-deoxyadenosine 5'-diphosphate (dADP3-) and 2'-deoxyadenosine 5'-triphosphate (dATP4-). These results are compared with previous data measured under the same conditions and available in the literature for the adenosine 5'-phosphates, AMP(2-), ADP(3-) and ATP(4-), as well as guanosine 5'-monophosphate (GMP(2-)) and 2'-deoxyguanosine 5'-monophosphate (dGMP(2-)). Hence, in total four nucleotide pairs, GMP(2-)/dGMP(2-), AMP(2-)/dAMP(2-), ADP(3-)/dADP(3-) and ATP(4-)/dATP(4-) (= NP/dNP), could be compared for the four metal ions Mg2+, Ni2+, Cu2+ and Zn2+ (= M2+). The comparisons show that complex stability and extent of macrochelate formation between the phosphate-coordinated metal ion and N7 of the purine residue is very similar (or even identical) for the AMP(2-)/dAMP(2-) and ADP(3-)/dADP(3-) pairs. In the case of the complexes formed with ATP(4-)/dATP(4-) the 2'-deoxy complexes are somewhat more stable and show also a slightly enhanced tendency for macrochelate formation. This is different for guanine nucleotides: the stabilities of the M(dGMP) complexes are clearly higher, as are the formation degrees of their macrochelates, than is the case with the M(GMP) complexes. This enhanced complex stability and greater tendency to form macrochelates can be attributed to the enhanced basicity (DeltapKaca. 0.2) of N7 in the 2'-deoxy compound. These results allow general conclusions regarding nucleic acids to be made. Among the 2'-deoxyribose nucleotides (dNMP 2-, dNDP 3-, dNTP 4-) the complexes of dGMP 2-and to a smaller extent also of dATP 4-are somewhat more stable than the corresponding ribose nucleotides (NMP 2-, NDP 3-, NTP 4-) and macrochelate formation involving N7 is also more pronounced. 3 SummaryThe interaction between metal ions and nucleotides is well characterized, as is their importance for metabolic processes, e.g., in the synthesis of nucleic acids. Hence, it is surprising to find that no detailed comparison is available of the metal ion-binding properties between nucleoside 5'-phosphates and 2'-deoxynucleoside 5'-phosphates. Therefore, we have measured here by potentiometric pH titrations the stabilities of several metal ion complexes formed with 2'-deoxyadenosine 5'-monophosphate (dAMP 2-), 2'-deoxyadenosine 5'-diphosphate (dADP 3-) and 2'-deoxyadenosine 5'-triphosphate (dATP 4-). These results are compared with previous data measured under the same conditions and available in the literature for the adenosine 5'-phosphates, AMP 2-, ADP 3-and ATP 4-as well as guanosine 5'-monophosphate (GMP 2-...

With a view on protein-nucleic acid interactions in the presence of metal ions we studied the "simple" mixed-ligand model systems containing histamine (Ha), the metal ions Ni(2+), Cu(2+), or Zn(2+) (M(2+)), and the nucleotides adenosine 5'-triphosphate (ATP(4-)) or uridine 5'-triphosphate (UTP(4-)), which will both be referred to as nucleoside 5'-triphosphate (NTP(4-)). The stability constants of the ternary M(NTP)(Ha)(2-) complexes were determined in aqueous solution by potentiometric pH titrations. We show for both ternary-complex types, M(ATP)(Ha)(2-) and M(UTP)(Ha)(2-), that intramolecular stacking between the nucleobase and the imidazole residue occurs and that the stacking intensity is approximately the same for a given M(2+) in both types of complexes: The formation degree of the intramolecular stacks is estimated to be 20 to 50%. Consequently, in protein-nucleic acid interactions imidazole-nucleobase stacks may well be of relevance. Furthermore, the well-known formation of macrochelates in binary M(2+) complexes of purine nucleotides, that is, the phosphate-coordinated M(2+) interacts with N7, is confirmed for the M(ATP)(2-) complexes. It is concluded that upon formation of the mixed-ligand complexes the M(2+)-N7 bond is broken and the energy needed for this process corresponds to the stability differences determined for the M(UTP)(Ha)(2-) and M(ATP)(Ha)(2-) complexes. It is, therefore, possible to calculate from these stability differences of the ternary complexes the formation degrees of the binary macrochelates: The closed forms amount to (65±10)%, (75±8)%, and (31±14) % for Ni(ATP)(2-), Cu(ATP)(2-), and Zn(ATP)(2-), respectively, and these percentages agree excellently with previous results obtained by different methods, confirming thus the internal validity of the data and the arguments used in the evaluation processes. Based on the overall results it is suggested that M(ATP)(2-) species, when bound to an enzyme, may exist in a closed macrochelated form only, if no enzyme groups coordinate directly to the metal ion.

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Interactions between sinefungin and copper(II) ions were investigated. Stoichiometry and stability constants of the metal-free system and two mononuclear complexes present in solution were determined on the basis of potentiometric data analysis. The results were compared to the Cu(II)-ornithine system due to structural similarities between both molecules. Combined spectroscopic and theoretical studies allowed for determination of coordination pattern for the Cu(II)-sinefungin complexes. At acidic pH, copper is bound in “glycine-like” coordination mode, identical with that of ornithine. This involves α-amino group and the carboxyl oxygen. At higher pH, a “bis-complex” is formed by two sinefungin molecules. The second ligand binds in equatorial position displacing two water molecules, what results in the stable {2N,2O} coordination. Both axial positions are supposed to be occupied by N1 nitrogen donors of adenine moiety, what is confirmed by DFT calculations. They interact indirectly with copper(II) through water molecules as the result of dominant syn conformation of purine.

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