Isothermal titration calorimetry reveals a zinc ion as an atomic switch in the diadenosine polyphosphates.
Diadenosine polyphosphates (diadenosine 5',5'''-P(1),P(n)-polyphosphate (Ap(n)A)) are 5'-5'''-phosphate-bridged dinucleosides that have been proposed to act as signaling molecules in a variety of biological systems. Isothermal titration calorimetry was used to measure the affinities of a variety of metal cations for ATP, diadenosine 5',5'''-P(1),P(3)-triphosphate (Ap(3)A), diadenosine 5',5'''-P(1),P(4)-tetraphosphate (Ap(4)A), and diadenosine 5',5'''-P(1),P(5)-pentaphosphate (Ap(5)A). The binding of Mg(2+), Ca(2+), and Mn(2+) to ATP is shown to take place with the beta,gamma-phosphates (primary site) and be endothermic in character. The binding of Ni(2+), Cd(2+), and Zn(2+) to ATP is found to take place at both the primary site and at a secondary site identified as N-7 of the adenine ring. Binding to this second site is exothermic in character. Generally, the binding of metal cations to diadenosine polyphosphates involves a similar primary site to ATP. No exothermic binding events are identified. Critically, the binding of Zn(2+) to diadenosine polyphosphates proves to be exceptional. This appears to involve a very high affinity association involving the N-7 atoms of both adenine rings in each Ap(n)A, as well as the more usual endothermic association with the phosphate chain. The high affinity association is also endothermic in character. A combination of NMR and CD evidence is provided in support of the calorimetry data demonstrating chemical shift changes and base stacking disruptions entirely consistent with N-7 bridging interactions. N-7 bridging interactions are entirely reversible, as demonstrated by EDTA titration. Considering the effects of Zn(2+) on a wide variety of dinucleoside polyphosphate-metabolizing enzymes, we examine the possibility of Zn(2+) acting as an atomic switch to control the biological function of the diadenosine polyphosphates.
Tanner
Julian A
JA
Imperial College Genetic Therapies Centre, Department of Chemistry, Imperial College of Science, Technology and Medicine, South Kensington, London SW7 2AZ, United Kingdom.
Abowath
Asif
A
Miller
Andrew D
AD
D000255:Adenosine Triphosphate / Q000737:chemistry; D001665:Binding Sites; D002151:Calorimetry / Q000379:methods; D002413:Cations, Divalent / Q000737:chemistry; D002942:Circular Dichroism; D015226:Dinucleoside Phosphates / Q000737:chemistry*; D008345:Manganese / Q000737:chemistry; D008968:Molecular Conformation; D015032:Zinc / Q000737:chemistry*
D002413:Cations, Divalent; D015226:Dinucleoside Phosphates; D008345:Manganese; C012275:P(1),P(5)-di(adenosine-5'-)pentaphosphate; C011440:diadenosine triphosphate; C070070:diadenosine 5',5''''-P1,P6-hexaphosphate; D000255:Adenosine Triphosphate; D015032:Zinc