Comparison of the metal-ion-promoted dephosphorylation of the 5'-triphosphates of adenosine, inosine, guanosine and cytidine by Mn2+, Ni2+ and Zn2+ in binary and ternary complexes.
The dependence of the rate of dephosphorylation of ATP, ITP, GTP and CTP (= NTP), expressed as first-order rate constants (50 degrees C; I = 0.1 M, NaClO4), on pH (2 to 10), in the absence and presence of Mn2+, Ni2+, and Zn2+, was investigated. The reaction is accelerated by Zn2+ and passes through a pH optimum at about 8 for the system Zn2+-ATP or 9 for Zn2+-ITP and Zn2+-GTP; this is analogous to observations made earlier with the corresponding Cu2+ systems. By computing the pH dependence of the distribution of the several species present in these systems it is shown that the highest rates are observed in the pH regions where the concentration of Zn(ATP)2-, Zn(ITP-H)3-, or Zn(GTP-H)3- dominates. By evaluating the pH dependence evidence is given that the attacking nucleophile is OH- or H2O for Zn (ATP)2- and H2O for Zn (ITP-H)3- or Zn(GTP-H)3-. For all these complexes metal-ion/nucleic-base interactions are known, leading to the formation of macrochelates. These metal-ion/nucleic-base interactions are crucial for the observation of a metal-ion-promoted dephosphorylation; in agreement with this, and the small tendency of the cytosine moiety to coordinate, the CTP systems are rather stable towards dephosphorylation. It should be noted that these experimental results do not necessarily mean that the macrochelates usually described are the reactive complexes, but only that the active complex must be closely related to them (e.g. isomers, etc). Although for the Ni2+ systems with ATP, ITP, and GTP, and for the Mn2+-ATP system a metal-ion/nucleic-base interaction is also known, these systems are not very sensitive to hydrolytic cleavage of the terminal P-O-P bond. The only known significant structural difference between the Ni2+-NTP or the Mn2+-ATP complexes and those of Cu2+ or Zn2+ is that Ni2+ Mn2+ coordinate to all three phsophate groups, whereas Cu2+ and Zn2+ involve only the beta and gamma ones. This structure-reactivity relationship is rationalized by the suggestion that in the active species the metal ion should be coordinated to the alpha,beta-phosphate groups leaving the gamma-group open to nucleophilic attack. Obviously, an initial beta,gamma-coordination is suitable for a shift of the metal ion along the phosphate back-bone into the reactive alpha-beta-position, while for an alpha,beta,gamma-coordination only the less favorable removal of the coordinated gamma-group remains. The metal-ion/nucleic-base interaction is considered as being important for achieving this reactive structure. The connection between trans-phosphorylation in vitro and in vivo is discussed. It is also shown that the formation of mixed-ligand or ternary complexes inhibits the dephosphorylation process. This is on the one hand of interest with regard to the transport of hydrolysis-sensitive phosphates in nature, while on the other it casts doubts on conclusions based on experiments carried out in the presence of buffers, because these contain weak bases and hence potential ligands.
Comparison of the metal-ion-promoted dephosphorylation of the 5'-triphosphates of adenosine, inosine, guanosine and cytidine by Mn2+, Ni2+ and Zn2+ in binary and ternary complexes. The dependence of the rate of dephosphorylation of ATP, ITP, GTP and CTP (= NTP), expressed as first-order rate constants (50 degrees C; I = 0.1 M, NaClO4), on pH (2 to 10), in the absence and presence of Mn2+, Ni2+, and Zn2+, was investigated. The reaction is accelerated by Zn2+ and passes through a pH optimum at about 8 for the system Zn2+-ATP or 9 for Zn2+-ITP and Zn2+-GTP; this is analogous to observations made earlier with the corresponding Cu2+ systems. By computing the pH dependence of the distribution of the several species present in these systems it is shown that the highest rates are observed in the pH regions where the concentration of Zn(ATP)2-, Zn(ITP-H)3-, or Zn(GTP-H)3- dominates. By evaluating the pH dependence evidence is given that the attacking nucleophile is OH- or H2O for Zn (ATP)2- and H2O for Zn (ITP-H)3- or Zn(GTP-H)3-. For all these complexes metal-ion/nucleic-base interactions are known, leading to the formation of macrochelates. These metal-ion/nucleic-base interactions are crucial for the observation of a metal-ion-promoted dephosphorylation; in agreement with this, and the small tendency of the cytosine moiety to coordinate, the CTP systems are rather stable towards dephosphorylation. It should be noted that these experimental results do not necessarily mean that the macrochelates usually described are the reactive complexes, but only that the active complex must be closely related to them (e.g. isomers, etc). Although for the Ni2+ systems with ATP, ITP, and GTP, and for the Mn2+-ATP system a metal-ion/nucleic-base interaction is also known, these systems are not very sensitive to hydrolytic cleavage of the terminal P-O-P bond. The only known significant structural difference between the Ni2+-NTP or the Mn2+-ATP complexes and those of Cu2+ or Zn2+ is that Ni2+ Mn2+ coordinate to all three phsophate groups, whereas Cu2+ and Zn2+ involve only the beta and gamma ones. This structure-reactivity relationship is rationalized by the suggestion that in the active species the metal ion should be coordinated to the alpha,beta-phosphate groups leaving the gamma-group open to nucleophilic attack. Obviously, an initial beta,gamma-coordination is suitable for a shift of the metal ion along the phosphate back-bone into the reactive alpha-beta-position, while for an alpha,beta,gamma-coordination only the less favorable removal of the coordinated gamma-group remains. The metal-ion/nucleic-base interaction is considered as being important for achieving this reactive structure. The connection between trans-phosphorylation in vitro and in vivo is discussed. It is also shown that the formation of mixed-ligand or ternary complexes inhibits the dephosphorylation process. This is on the one hand of interest with regard to the transport of hydrolysis-sensitive phosphates in nature, while on the other it casts doubts on conclusions based on experiments carried out in the presence of buffers, because these contain weak bases and hence potential ligands.