Neu Ancilla, Neu Ursula, Fuchs Anna-Lisa, Schlager Benjamin, Sprangers Remco
Max Planck Institute for Developmental Biology, Tübingen, Germany.
Interfaculty Institute of Biochemistry, University of Tübingen, Tübingen, Germany.
Nat Chem Biol. 2015 Sep;11(9):697-704. doi: 10.1038/nchembio.1866. Epub 2015 Aug 10.
The scavenger decapping enzyme hydrolyzes the protective 5' cap structure on short mRNA fragments that are generated from the exosomal degradation of mRNAs. From static crystal structures and NMR data, it is apparent that the dimeric enzyme has to undergo large structural changes to bind its substrate in a catalytically competent conformation. Here we studied the yeast enzyme and showed that the associated opening and closing motions can be orders of magnitude faster than the catalytic turnover rate. This excess of motion is induced by the binding of a second ligand to the enzyme, which occurs at high substrate concentrations. We designed a mutant that disrupted the allosteric pathway that links the second binding event to the dynamics and showed that this mutant enzyme is hyperactive. Our data reveal a unique mechanism of substrate inhibition in which motions that are required for catalytic activity also inhibit efficient turnover when they are present in excess.
清道夫去帽酶可水解短mRNA片段上的保护性5'帽结构，这些短mRNA片段是由mRNA的外泌体降解产生的。从静态晶体结构和核磁共振数据来看，很明显二聚体酶必须经历大的结构变化，才能以催化活性构象结合其底物。在这里，我们研究了酵母酶，结果表明相关的打开和关闭运动可能比催化周转速度快几个数量级。这种过量的运动是由第二种配体与酶的结合诱导的，这种结合发生在高底物浓度下。我们设计了一个突变体，破坏了将第二次结合事件与动力学联系起来的变构途径，结果表明这种突变酶具有高活性。我们的数据揭示了一种独特的底物抑制机制，即催化活性所需的运动在过量存在时也会抑制有效周转。
