Abstract
Replacement of a non‐bridging oxygen in the phosphate diester bond by a sulfur has become quite popular in nucleic acid research and is often used as a probe, for example, in ribozymes, where the normally essential Mg^2+^ is partly replaced by a thiophilic metal ion to reactivate the system. Despite these widely applied rescue experiments no detailed studies exist quantifying the affinity of metal ions to such terminal sulfur atoms. Therefore, we performed potentiometric pH titrations to determine the binding properties of pUp~(S)~U^3−^ towards Mg^2+^, Mn^2+^, Zn^2+^, Cd^2+^, and Pb^2+^, and compared these data with those previously obtained for the corresponding pUpU^3−^ complexes. The primary binding site in both dinucleotides is the terminal phosphate group. Theoretically, also the formation of 10‐membered chelates involving the terminal oxygen or sulfur atoms of the (thio)phosphate bridge is possible with both ligands. The results show that Mg^2+^ and Mn^2+^ exist as open (op) isomers binding to both dinucleotides only at the terminal phosphate group. Whereas Cd(pUpU)^−^ only exists as Cd(pUpU)${{{- \hfill \atop {\rm op}\hfill}}}$, Cd(pUp~(S)~U)^−^ is present to about 64 % as the S‐coordinated macrochelate, Cd(pUp~(S)~U)${{{- \hfill \atop {\rm cl}/{\rm PS}\hfill}}}$. Zn^2+^ forms with pUp~(S)~U^3−^ three isomeric species, that is, Zn(pUp~(S)~U)${{{- \hfill \atop {\rm op}\hfill}}}$, Zn(pUp~(S)~U)${{{- \hfill \atop {\rm cl}/{\rm PO}\hfill}}}$, and Zn(pUp~(S)~U)${{{- \hfill \atop {\rm cl}/{\rm PS}\hfill}}}$, which occur to about 33, 12 (O‐bound), and 55 %, respectively. Pb^2+^ forms the 10‐membered chelate with both nucleotides involving only the terminal oxygen atoms of the (thio)phosphate bridge, that is, no indication of S binding was discovered in this case. Hence, Zn^2+^ and Cd^2+^ show pronounced thiophilic properties, whereas Mg^2+^, Mn^2+^, and Pb^2+^ coordinate to the oxygen, macrochelate formation being of relevance with Pb^2+^ only.