Krapp Stephan, Münster-Kühnel Anja K, Kaiser Jens T, Huber Robert, Tiralongo Joe, Gerardy-Schahn Rita, Jacob Uwe
Max-Planck-Institut für Biochemie, Abteilung für Strukturforschung, Am Klopferspitz 18a, 82152, Martinsried, Germany.
J Mol Biol. 2003 Dec 5;334(4):625-37. doi: 10.1016/j.jmb.2003.09.080.
Sialic acids are activated by CMP-5-N-acetylneuraminic acid synthetase prior to their transfer onto oligo- or polysaccharides. Here, we present the crystal structure of the N-terminal catalytically active domain of the murine 5-N-acetylneuraminic acid synthetase in complex with the reaction product. In contrast to the previously solved structure of 5-N-acetylneuraminic acid synthetase from Neisseria meningitidis and the related CMP-KDO-synthetase of Escherichia coli, the murine enzyme is a tetramer, which was observed with the active sites closed. In this conformation a loop is shifted by 6A towards the active site and thus an essential arginine residue can participate in catalysis. Furthermore, a network of intermolecular salt-bridges and hydrogen bonds in the dimer as well as hydrophobic interfaces between two dimers indicate a cooperative behaviour of the enzyme. In addition, a complex regulation of the enzyme activity is proposed that includes phosphorylation and dephosphorylation.
唾液酸在转移到寡糖或多糖之前，由CMP-5-N-乙酰神经氨酸合成酶激活。在此，我们展示了与反应产物复合的小鼠5-N-乙酰神经氨酸合成酶N端催化活性结构域的晶体结构。与先前解析的脑膜炎奈瑟菌5-N-乙酰神经氨酸合成酶以及大肠杆菌相关的CMP-KDO合成酶的结构不同，小鼠酶是四聚体，观察到其活性位点是关闭的。在这种构象中，一个环向活性位点移动了6埃，因此一个必需的精氨酸残基可以参与催化作用。此外，二聚体中的分子间盐桥和氢键网络以及两个二聚体之间的疏水界面表明该酶具有协同行为。另外，还提出了一种包括磷酸化和去磷酸化的酶活性复杂调节机制。
