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

Abstract

The dependence of the rate of dephosphorylation of ATP, ITP, GTP and CTP (= NTP), expressed as first-order rate constants (50 "C; I = 0.1 M, NaCIO,), on pH (2 to lo), in the absence and presence of Mn2+, Ni2+ and Zn2+, was investigated. The reaction is accelerated by Zd2+ and passes through a pH optimum at about 8 for the system Zn2f.-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(lTP-H)3-, or Zn(GTP-H)3 -dominates. By evaluating the pH dependence evidence is given that the attacking nucleophije is OH-or H 2 0 for Zn(ATP)'-and H20 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-0-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+ and Mn2+ coordinate to all three phosphate groups, whereas Cu2 + and Zn2 + involve only the fl and y ones. This structure-reactivity relationship is rationalized by the suggestion that in the active species the metal ion should be coordinated to the a,/?-phosphate groups leaving the 1'-group open to nucleophilic attack. Obviously, an initial PJ-COordination is suitable for a shift of the metal ion along the phosphate back-bone into the reactive a,P-position, while for an a,/?,?-coordination only the less favorable removal of the coordinated pgroup remains. The metal-ion/nucleic-base interaction is considered as being important for achieving this reactive structure. The connection between trans-phosphorylations 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.Considering the importance of phosphoryl and nucleotidyl transfers in nature and because the enzymes ...

Properties of ribose modified ADP analogues in photophosphorylation of spinach chloroplasts

“…Column: 300 X 5 mm; NucleosilR 5 SB, silica gel spherical TP, particle size 5 firn (Machery & Nagel, Diiren Germany); mobile phase: 0.1 N KNO,, 0.02 M KH,PO, pH 2.6; flux rate: 21 ml/h, 67 atm, 24°C; detection system: Zeiss PMQIV, 258 nm. nated directly or via water molecules to the CY-and P-phosphoryl group in the active complex [27]. This stereo-specific Mg"-ADP complex seems to be necessary for a proper positioning of the nucleotide after binding to the active site.…”

Metal‐Ion‐Promoted Dephosphorylation of the 5′ ‐Triphosphates of Uridine and Thymidine, and a Comparison with the Reactivity in the Corresponding Cytidine and Adenosine Nucleotide Systems

“…S3A and S3B). Using the mentioned arguments (see Supplenicnt) we postulate again that the reactive species are the isocharged complexes Zn(UTP)(OH)3 -and Ni(UTP)(OH)3 -, as was concluded earlier [7] for Zn(CTP)(OH)3-and Ni(CTP)(OH)3 -.…”

“…The corresponding pH,, was obtained by averaging the pH values measured for those samples that gave points on the straight-line portion. Examples are shown in the figures of [7] and [14].…”

“…It is known that different metal ions promote the selfassociation differently [6] ; the ' extent of macrochelate formation in the monomeric complexes by the coordination of a metal ion to the phosphate chain and to N-7 is also different [6]. All this is reflected in the varying effectiveness of metal ions in promoting the dephosphorylation of ATP [ 1,7]. This is Part VII of the series Hydrolysis of Nucleoside Phosphutes.…”

Characterization of Metal Ion-Nucleic Acid Interactions in Solution

“…[307]. Phosphate, which is frequently employed as a buffer in spectroscopic or biochemical experiments of nucleic acids, also interacts with metal ions strongly enough to falsify measurements of intrinsic affinity constants [36] or of reaction rates [312][313][314][315], an observation that holds for other buffers as well [312,316]. Consequently, one is often forced to work in buffer-free media [312,317,318].…”