The chemical shift of the protons, H(2) and H(8), in the proton magnetic resonance spectra of ATP, ITP, and inosine was measured in aqueous solution in dependence on pH (2 to 12). Based on these data the sites of protonation or deprotonation are identified and the acidity constants determined. A comparison of the constants due to the nucleotides with the corresponding ones of the nucleosides evidences that the basicity of the nucleic base increases somewhat under the influence of the triphosphate chain. The possibility of an intramolecular hydrogen bound between N(7) and the proton of the y-phosphate group is discussed.From the line broadening effects in the presence of Cu2+ it is concluded that this metal ion interacts mainly with N(7) of ATP, but with the N(l), 0 (6) is nearly completely formed) reveal the surprising fact that in the pH range 2 to 6 the line broadening of the signal due to H(8), and H(2) as well, increases signiticantly compared with a system containing only ITP and Cua+. This observation is easily explained by the assumption of a chargetransfer interaction within the ternary complex, Cu(Bipy) (ITP)2-, i.e. between the purine part of the nucleotide and 2,2'-bipyridyl. This interaction could indeed be contirmed by spectrophotometric measurements in the ultraviolet (Amax = 313 nm). Similar results were obtained for the mixed-ligand complex, Cu(Bipy) (ATP)2-. The distance between the planes of the aromatic rings in these systems is estimated and a structure for these metal-ion-bridged charge-transfer complexes is suggested. The biological implications of such ternary, metal-ion-bridged, charge-transfer complexes are briefly indicated.Recent studies on the dephosphorylation of the binary and ternary complexes, CU(ATP)~-and Cu(Bipy)(ATP)2- [3,4], and the extension of this work to other nucleotide complexes (H. Sigel, D. H. Buisson and B. Prijs, unpublished results) prompted us to investigate the coordination behavior of ITP in mixed-ligand complexes. For reasons of comparison inosine and ATP were included' in this work. From earlier studies on binary complexes [6,6] it is known that Cu2+ coordinates to the j3-and y-phosphate groups of ATP4-as well as to N(7) of the adenine moiety, and thus a macrochelate is formed [6, 71, which is crucial indeed for the Cu2+-accelerated dephosphorylation of ATP [2,4,7]. Unfortunately, the coordination behavior of ITP is less clear cut. However, as the N(7); usually the latter two are favored.In solution, Cu2+ prefers a square-planar (or grossly distorted) coordination sphere [13,14] ; hence, in Cu(Bipy)2+ only two positions remain available for the binding of a second ligand [la], the coordination of which results in a ternary complex. In fact, for Cu(Bipy)(ATP)2-it was shown by means of proton magnetic resonance (PMR) that in this mixedligand complex the adenine moiety does not interact any more with Cu2+ [16]. It is believed that this is Eur. J. Biochem. 41 (1974)
