Scott W G, Murray J B, Arnold J R, Stoddard B L, Klug A
MRC Laboratory of Molecular Biology, Hills Road, Cambridge CB2 2QH, England.
Science. 1996 Dec 20;274(5295):2065-9. doi: 10.1126/science.274.5295.2065.
The crystal structure of an unmodified hammerhead RNA in the absence of divalent metal ions has been solved, and it was shown that this ribozyme can cleave itself in the crystal when divalent metal ions are added. This biologically active RNA fold is the same as that found previously for two modified hammerhead ribozymes. Addition of divalent cations at low pH makes it possible to capture the uncleaved RNA in metal-bound form. A conformational intermediate, having an additional Mg(II) bound to the cleavage-site phosphate, was captured by freeze-trapping the RNA at an active pH prior to cleavage. The most significant conformational changes were limited to the active site of the ribozyme, and the changed conformation requires only small additional movements to reach a proposed transition-state.
在不存在二价金属离子的情况下，未修饰的锤头状RNA的晶体结构已得到解析，结果表明，当添加二价金属离子时，这种核酶在晶体中能够自我切割。这种具有生物活性的RNA折叠结构与之前发现的两种修饰的锤头状核酶相同。在低pH值下添加二价阳离子，使得能够捕获以金属结合形式存在的未切割RNA。通过在切割前将RNA在活性pH值下进行冷冻捕获，捕捉到了一种构象中间体，该中间体在切割位点的磷酸上额外结合了一个Mg(II)。最显著的构象变化仅限于核酶的活性位点，而这种变化后的构象只需进行少量额外移动就能达到假定的过渡态。
