Nuclear Magnetic Resonance Spectra of Adenosine Di- and Triphosphate II. EFFECT OF COMPLEXING WITH DIVALENT METAL IONS*

The requirement of a divalent metal ion for all enzymatic reactions involving adenosine diand triphosphate is well established, but the role of the metal ion is not clearly understood. In some kinase reactions (1, 2) kinetic evidence has been presented that the metal complex of adenosine triphosphate is the true substrate in the enzymatic reaction. The present study was undertaken to obtain information concerning changes in the structure of adenosine diand triphosphate caused by interactions with various divalent metal ions. Such information might shed some light on the changes in the reactivity of specific bonds in the nucleotides induced by the met’al ions and would certainly impose limitations on any proposed structure of an enzyme-substrate complex for these compounds. In a previous paper (3), it was shown that chemical shifts of the nuclear magnetic resonance peaks of the phosphorus nuclei of adenosine diand triphosphate could be measured on 0.1 M solutions of the nucleotides. The same techniques have been applied in the present investigation to observe the changes in the chemical shifts of the phosphorus magnetic resonance spectra in the presence of metal ions to give direct evidence of the nature of the complexes formed. With the use of high resolution technique, the chemical shifts of the proton magnetic resonance spectra in the nucleotides could be compared with the shifts in the metal complexes to determine the interaction of the metal ion with the adenine ring. The measurements were extended to include the effect of paramagnetic ions such as Mn ++ and Cu++ at low concentrations of the order of 1O-4 to 1O-5 M on the width of the absorption lines of both phosphorus and protons. Such spectra revealed preferential interactions with particular phosphorus nuclei and protons, thus specifying the nature of the met.al complexes formed.