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The acidity constants of guanylyl(3'-->5')guanosine (GpG(-)) and 2'-deoxyguanylyl(3'-->5')-2'-deoxyguanosine [d(GpG)(-)] for the deprotonation of their (N1)H sites were measured by potentiometric pH titrations in aqueous solution (25 degrees C; I = 0.1 M, NaNO(3)). The same method was used for the determination of the stability constants of the 1:1 complexes formed between Mg(2+), Ni(2+), or Cd(2+) (= M(2+)) and (GG-H)(2-), and in the case of Mg(2+) also of (GG-2H)(3-), where GG(-) = GpG(-) or d(GpG)(-). The stability constants of the M(GG)(+) complexes were estimated. The acidity constants of the H(dGuo)(+) and dGuo species (dGuo = 2'-deoxyguanosine) and the stability constants of the corresponding M(dGuo)(2+) and M(dGuo-H)(+) complexes were also measured. Comparison of these and related data allows the conclusion that N7 of the 5'G unit in GG(-) is somewhat more basic than the one in the 3'G moiety; the same holds for the (N1)(-) sites. On the basis of comparisons with the stability constants measured for the dGuo complexes, it is concluded that M(2+) binding of the GG dinucleoside monophosphates occurs predominantly in a mono-site fashion, meaning that macrochelate formation is not very pronounced. Indeed, it was a surprise to find that the stabilities of the complexes of dGuo or (dGuo-H)(-) and the corresponding ones derived from GG(-) are so similar. Consequently, it is suggested that in the M(GG)(+) and M(GG-H) complexes the metal ion is mainly located at N7 of the 5'G unit since this is the more basic site allowing also an outer-sphere interaction with the C6 carbonyl oxygen and because this coordination mode is also favorable for an electrostatic interaction with the negatively charged phosphodiester bridge. It is further suggested that Mg(2+) binding (which is rather weak compared to that of Ni(2+) and Cd(2+)) occurs mainly in an outer-sphere mode, and on the basis of the so-called Stability Ruler it is concluded that the binding properties of Zn(2+) to the GG species are similar to those of Ni(2+) and Cd(2+).

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The acidity constants of H 2 (AcP) and H 2 (AnP), where AcP 2-) acetyl phosphate and AnP 2-) acetonylphosphonate, as well as the stability constants of the 1:1 complexes formed between Mg 2+ , Ca 2+ , Sr 2+ , Ba 2+ , Mn 2+ , Co 2+ , Ni 2+ , Cu 2+ , Zn 2+ , or Cd 2+ and AcP 2-or AnP 2-were determined by potentiometric pH titrations in aqueous solution (25°C; I ) 0.1 M, NaNO 3 ). On the basis of previously established log K M(R-PO 3 ) M versus pK H(R-PO 3 ) H straight-line plots for phosph(on)ate ligands (R-PO 3 2-), which allow only a simple -PO 3 2-coordination (Sigel, H.; et al. HelV. Chim. Acta 1992, 75, 2634), it is concluded that the carbonyl oxygen atom of AcP 2-and AnP 2-participates in complex formation by giving rise to six-membered chelates for all of the mentioned systems except for Ba(AnP) and Sr(AnP), the stability of which corresponds to a simple -PO 3 2-coordination. The formation degree of the chelates formed by the alkaline earth ions and AcP 2-is quite pronounced; it amounts, for example, for Mg(AcP) and Ca(AcP) to 41% ( 5% and 48% ( 8%, respectively. The corresponding results for Zn(AcP) and Cu(AcP) are 59% ( 6% and 76% ( 4%, respectively. The formation degree of the six-membered Cu(AnP) chelate increases in water containing 30% or 50% (v/v) 1,4-dioxane. This may also be surmised for the other divalent metal ions under conditions of lower solvent polarity and poorer solvating properties than water. Such conditions exist in active-site cavities of enzymes, and the expected effects are shortly discussed. The indicated measurements with AcP have only become possible after the stabilities of the corresponding M(HPO 4 ) complexes had been determined (Saha, A.; et al. J. Biol. Inorg. Chem. 1996, 1, 231) because AcP contains as an impurity always some phosphate; this phosphate content had to be quantified and its effect was carefully considered in the evaluations of the experimental data. Here is most probably also the main reason why previously published results regarding the stabilities of the Mg(AcP) and Ca(AcP) complexes vary widely; stability constants of the other M(AcP) complexes have not been determined previously.

The acidity constants of twofold protonated methyl thiophosphate (MeOPS(2-)) and of monoprotonated uridine 5'- O-thiomonophosphate (UMPS(2-)) have been determined in aqueous solution (25 degrees C; I= 0.1 M, NaNO(3)) by potentiometric pH titration. The stability constants of their 1:1 complexes formed with Pb(2+), i.e. Pb(MeOPS) and Pb(UMPS), have also been measured. The results show that replacement of a phosphate oxygen by a sulfur atom increases the acidity by about 1.4 p K units. On the basis of recently established log versus plots ( = simple phosphate or phosphonate ligands where R is a non-coordinating residue), it is shown that the stability of the Pb(thiophosphate) complexes is by log Delta= 2.43+/-0.09 larger than expected for a Pb(2+)-phosphate interaction. The identity of the stability increase (log Delta) observed for Pb(MeOPS) and Pb(UMPS) shows that the nucleobase residue in the Pb(UMPS) complex has no influence on complex formation. To be able to carry out the mentioned comparisons, we have also determined the stability constant of the complex formed between Pb(2+) and methyl phosphate; the corresponding data for Pb(UMP) were already known from our earlier studies. The present results allow an evaluation of other Pb(2+) complexes formed with thiophosphate derivatives and they are applied now to the Pb(2+) complexes of adenosine 5'- O-thiomonophosphate (AMPS(2-)). The stability constants of the Pb(H;AMPS)(+) and Pb(AMPS) complexes were measured and it is shown that, within the error limits, the stability of the Pb(AMPS) complex is determined by the basicity of the thiophosphate group of AMPS(2-); in other words, no hint for macrochelate formation involving N7 was observed. More important, with the aid of micro-stability-constant considerations it is concluded that the structure of the dominating isomer of the Pb(H;AMPS)(+) species is the one where the proton is located at the N1 site of the adenine residue and Pb(2+) is coordinated to the deprotonated thiophosphate group. The insights gained from this study with regard to thiophosphate-altered single-stranded nucleic acids and their affinity towards Pb(2+) are discussed.

The stability constants of the 1:1 complexes formed in aqueous solution between Mg2+, Ca2+, Sr2+, Ba2+, Mn2+, Co2+, Ni2+, Zn2+, or Cd2+ (M2+) and methyl thiophosphate (MeOPS(2-)) or uridine 5'-O-thiomonophosphate (UMPS(2-)) (PS(2-)=MeOPS(2-) or UMPS(2-)) have been determined (potentiometric pH titrations; 25 degrees C; I = 0.1 M, NaNO(3)). Comparison of these results for M(PS) complexes with those known for the parent M(PO) phosphate species, where PO(2-)=CH(3)OPO(2-)(3) or UMP(2-) (uridine 5'-monophosphate), shows that the alkaline earth metal ions, as well as Mn2+, Co2+, and Ni2+ have a higher affinity for phosphate groups than for their thio analogues. However, based on the linear log K(M)(M(R-PO3)) versus pK(H)(H(R-PO3)) relationships (R-PO(2-)(3) simple phosphate monoester or phosphonate ligands with a non-interacting residue R) it becomes clear that the indicated observation is only the result of the lower basicity of the thiophosphate residue. In contrast, the thio complexes of Zn2+ and Cd2+ are more stable than their parent phosphate ones, and this despite the lower basicity of the PS(2-) ligands. This stability increase is identical for M(MeOPS) and M(UMPS) species and amounts to about 0.6 and 2.4 log units for Zn(PS) and Cd(PS), respectively. Since no other binding site is available in MeOPS(2-), this enhanced stability has to be attributed to the S atom. Indeed, from the mentioned stability differences it follows that Cd2+ in Cd(PS) is coordinated by more than 99% to the thiophosphate S atom; the same value holds for Pb(PS), which was studied earlier. The formation degree of the Sbonded isomer amounts to 76+/-6 % for Zn(PS) and is close to zero for the corresponding Mg2+, Ca2+, and Mn2+ species. It is further shown that Zn(MeOPS)(aq)(2+) releases a proton from a coordinated water molecule with pK(a) approximately 6.9; i.e., this deprotonation occurs at a lower pH value than that for the same reaction in Zn(aq)(2+). Since Mg2+, Ca2+, Mn2+, and Cd2+ have a relatively low tendency for hydroxo complex formation, it was possible, for these M2+, to also quantify the stability of the binuclear complexes, M(2)(UMPS-H)+, where one M2+ is thiophosphate-coordinated and the other is coordinated at (N3)(-) of the uracil residue. The impact of the results presented herein regarding M2+/nucleic acid interactions, including those of ribozymes (rescue experiments), is briefly discussed.

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