Joseph Granot

On the role of ATP and divalent metal ions in the storage of catecholamines

The storage vesicles of catecholamines in the adrenal medulla, i.e., the chromaffln granules, contain a remarkably high content of catecholamine (> 0.5 M) and nucleotides (mainly ATP, -0.1 M), as well as substantial amounts of acidic protein (chromogranin A) and divalent metal ions [ 1,2]. The mechanism by which such high concentrations are stored is unknown. Suggestions have been made that catecholamines participate in non-diffusible storage complexes with ATP [3,4] or with ATP together with divalent metal ions [S], or that catecholamines form aggregates with ATP which may play a role in their storage [6]. The possible formation of either binary catecholamineATP complexes or ternary complexes with metal ions, in aqueous solution, has been demonstrated by several spectroscopic studies [7-l 51. Recently, on the basis of potentiometric and ultraviolet absorption measurements of ternary catecholamine-metal ion-ATP complexes, a ‘metal-coordination hypothesis’ [ 161 was proposed in which the storage of catecholamine is attributed to the formation of multinuclear structures which contain catecholamine, ATP, divalent metal ions, and possibly phospholipids. It suggests that catecholamine chelate divalent metal ions either through ionized ring hydroxyls or through the amine and the &hydroxyl of the side chain. However, examination of this hypothesis reveals several incon-

Effect of chemical exchange on the transverse relaxation rate of nuclei in solution containing paramagnetic ions

Abstract An analysis of the exchange contribution to the transverse relaxation rate of nuclei in bulk solvent molecules, in a dilute solution containing paramagnetic ions, in limiting and intermediate cases, is presented. The conditions under which the absorption mode signal of these nuclei has a Lorentzian line shape are examined in detail. As a result, a constraint on the concentration of the solution under investigation is obtained.

Phosphorus-31 relaxation times of inorganic phosphate and nucleotides in aqueous solution

Abstract 31 P spin-lattice relaxation times ( T 1 ) of inorganic orthophosphate, and of the nucleotides TMP, AMP and ATP are reported as a function of pH. The T 1 values are found to exhibit significant pH dependences, with minimum T 1 values generally observed at pH values close to the p K a values for the orthophosphates. The effects of the addition of Cu 2+ or EDTA and treatment with cation exchange resin are described. These observations are related to similar results obtained previously for other ionizing species using 1 H and 13 C NMR, indicating a general phenomenon of such pH-dependent variations in T 1 values in aqueous solutions. It is suggested that this effect stems from interactions with paramagnetic metal ion impurities.

Proton magnetic resonance study of the dysprosium(III) aqueous complex

Abstract Proton field shifts in dysprosium perchlorate aqueous solutions were measured as function of temperature and dysprosium concentration. The contact and pseudo-contact shifts were separated by means of a least-squares method based on their different temperature variation (as T −1 and T −2 , respectively). The hyperfine coupling constant between the dysprosium unpaired electrons and the proton nuclei, and the spin density at the proton nuclei were calculated. The results were compared with those obtained for other lanthanides and interpreted in terms of structural changes. Linewidth measurements yielded an estimation for the water exchange rate. The electronic relaxation times were calculated and the activation energies for the relaxation processes present in the solution were obtained.

NMR studies of catecholamines. Interactions with adenine nucleotides by 31P magnetic resonance

Elucidation, on the molecular level, of the interactions between catecholamines and adenine nudeotides, particularly ATP, can provide a basis for understanding the biochemical mechanisms involved in the processes of storage, release, uptake and neural action of catecholamines. IH-NMR spectroscopy studies [ 1 ] have demonstrated that catecholamines and adenine nucleotides form in aqueous solution binary complexes primarily through stacking interaction between the catechol and the adenine rings. In addition, an electrostatic interaction between the positively charged ammonium group of the catecholamines and the negative phosphate moiety of the nucleotides was found to play an important role in stabilizing these complexes. It is the purpose of the present study to further investigate the involvement of the phosphate groups of adenIne nucleotides In the association with catecholamines, by means of 31P-NMR. Chemical shifts, spin couplings and spin-lattice relaxation times were measured for the phosphorus resonances of the nucleotides. The effects of catecholamines were found to be rather small, apparently due to the relatively weak, through-space, interactions involved. However, due to the high sensitivity of phosphate groups to their chemical environment, the results were of sufficient significance to allow gaining a better insight into the phosphate-amine interactions.