Investigation of the ternary D-myo-inositol 1,2,6-tris(phosphate)-spermine-Zn2+ system in solution.

Interactions of Ins(1,2,6)P3 (IP), with spermine (Spm) and zinc cations have been studied by potentiometric and 31P NMR titrations. In the 4-11 pH range, two IPSpmZn2H3 and IPSpmZn2H mixed complexes are formed which are largely predominant with respect to the binary species. According to 31P NMR titration it is likely that one of the zinc cations preferably binds phosphates P1 and P6. The adduct formation between Ins(1,2,6)P3 and spermine seems also favourable to the formation of the mixed complexes. The occurrence of ternary complexes involving inositol-phosphates, biogenic amines, and metallic cations may be of relevance in the regulation of biological processes.

Inframolecular studies of the protonation of adenophostin A: comparison with 1-D-myo-inositol 1,4,5-trisphosphate.

Adenophostin A is a glyconucleotide natural product with the highest known potency for the D-myo-inositol 1,4,5-trisphosphate receptor. Using synthetic adenophostin A we have investigated the macroscopic and microscopic protonation process of this compound by performing (31)P NMR, (1)H NMR, and potentiometric titration experiments. The logarithms of the first to the fourth stepwise protonation constants are, respectively, log K(1) = 8.48, log K(2) = 6.20, log K(3) = 4.96, and log K(4) = 3.80. The latter constant refers to the protonation equilibrium involving the N1 adenine nitrogen. From the microconstants the protonation fractions of each individual phosphate group can be calculated. Remarkably, the ionization state of the phosphates of adenophostin A at near physiological pH is very similar to those of inositol 1,4,5-trisphosphate, indicating that differences in phosphate charge cannot account for the high potency of this molecule. The analysis of the (1)H chemical shifts vs pH provided complementary conformational information. In particular, a slight "wrongway shift" of H1" can be related to the protonation of P2, thus indicating a short H1"-P2 distance. Our results are in line with a recently published model in which, however, a certain degree of constraint would keep the ribose 2'-phosphate moiety close to the glucose ring phosphates.