Hediger M, Milburn R M
J Inorg Biochem. 1982 Jun;16(3):165-82. doi: 10.1016/s0162-0134(00)80105-1.
Complex formation between ATP (adenosine 5'-triphosphate) and tn2COIII(aq) (tn = trimethylenediamine) and resulting hydrolysis of the ATP to ADP (adenosine 5'-diphosphate), AMP (adenosine 5'-monophosphate), PPi (pyrophosphate), and Pi (orthophosphate) have been examined by means of 31P nmr. With ATP approximately 0.1 M and tn2CoIII-(aq) up to 0.3 M, complex formation was promoted by equilibrating solutions for a period at pH 4, after which hydrolysis was allowed to proceed at each of several pHs in the range 5 to 9 prior to quenching by addition of strong base. With ATP 0.01 M and tn2CoIII(aq) up to 0.08 M, the above procedure was followed in some cases; in other experiments the pH of each ATP/tn2CoIII(aq) solution was adjusted immediately to a value in the range 5 to 9 with the remainder of the procedure as before. In most cases the hydrolysis was at 25 degrees C, but temperature dependence was also examined. The integrals for the beta-phosphorus resonance have been used to analyze for ATP in the quenched solutions; independent measurements of ATP by an enzyme/spectrophotometric method (Bergmeyer) gave similar results. Cobalt to ATP molar ratios up to 1 produce tn2CoIII-ATP as the predominant ATP complex; this 1:1 complex shows no detectable acceleration in hydrolysis compared to free ATP. Cobalt to ATP molar ratios of greater than 1 lead to complexes of type (tn2CoIII)2ATP and (tn2CoIII)3ATP, which exhibit greatly enhanced reactivity towards ATP hydrolysis. At a 2:1 molar ratio (0.1 or 0.01 M ATP), the enhancement in rate is approximately 10(5) at pH 7 where the rate is a maximum (comparison for 25 degrees C); at higher molar ratios the rate enhancements are even greater. The results support the view that effective metal ion catalysis of ATP hydrolysis requires formation of reactive species involving more than one metal ion per ATP.
通过³¹P核磁共振研究了ATP（腺苷5'-三磷酸）与tn2COIII(aq)（tn =三亚甲基二胺）之间的络合物形成以及由此导致的ATP水解为ADP（腺苷5'-二磷酸）、AMP（腺苷5'-单磷酸）、PPi（焦磷酸）和Pi（正磷酸）的过程。对于约0.1 M的ATP和高达0.3 M的tn2CoIII-(aq)，通过在pH 4下将溶液平衡一段时间来促进络合物形成，之后在加入强碱淬灭之前，让水解在5至9的几个pH值下进行。对于0.01 M的ATP和高达0.08 M的tn2CoIII(aq)，在某些情况下遵循上述步骤；在其他实验中，将每个ATP/tn2CoIII(aq)溶液的pH值立即调节至5至9的范围内，其余步骤如前。在大多数情况下，水解在25℃下进行，但也研究了温度依赖性。已使用β-磷共振的积分来分析淬灭溶液中的ATP；通过酶/分光光度法（Bergmeyer）对ATP进行的独立测量给出了类似的结果。钴与ATP的摩尔比高达1时，产生tn2CoIII-ATP作为主要的ATP络合物；与游离ATP相比，这种1:1络合物在水解中未显示出可检测到的加速作用。钴与ATP的摩尔比大于1时会导致(tn2CoIII)2ATP和(tn2CoIII)3ATP类型的络合物，它们对ATP水解表现出大大增强的反应性。在2:1摩尔比（0.1或0.01 M ATP）下，在速率最大的pH 7（25℃时的比较）时，速率增强约为10⁵；在更高的摩尔比下，速率增强更大。结果支持这样一种观点，即有效的金属离子催化ATP水解需要形成每个ATP涉及不止一个金属离子的反应性物种。
