Hohenester E, Keller J W, Jansonius J N
Department of Structural Biology, University of Basel, Switzerland.
Biochemistry. 1994 Nov 22;33(46):13561-70. doi: 10.1021/bi00250a008.
The pyridoxal 5'-phosphate-dependent enzyme dialkylglycine decarboxylase (DGD) is activated by K+ and Rb+ ions, whereas Li+ and Na+ ions are inhibitory. A binding site for alkali metal ions close to the active site (site 1) was discovered in the crystal structure of DGD, and an exchange of K+ for Na+ at this site was shown to affect the conformation of two active site residues [Toney, M. D., Hohenester, E., Cowan, S. W., & Jansonius, J. N. (1993) Science 261, 756-759]. We have investigated the effects of alkali metal ions on DGD activity and have determined the crystal structures at 2.8 A resolution of DGD with Li+ and Rb+ bound at site 1. Due to the weak scattering of the Li+ ion, its position had to be modeled using information from small molecule structures. A comparison of the DGD structures with Li+, Na+, K+, and Rb+ bound at site 1 reveals a striking correlation between active site structure and enzymatic activity. The small, inhibitory ions Li+ and Na+ are accommodated by replacing two protein-derived ligands of the larger, activating ions K+ and Rb+ by a single water molecule. This actuates a two-state structural switch between active and inactive enzyme that involves a concerted reorientation of the active site residues Ser80 and Tyr301 and a small change in the quaternary structure of the DGD tetramer. An important role of the essential K+ ion in both cofactor binding and the organization of a catalytically competent active site structure is proposed. In the structure of DGD with Rb+ bound at site 1, a second Rb+ ion has partially replaced the structural Na+ ion at metal binding site 2 on the surface of the DGD molecule, without significantly altering the protein structure. In contrast to Na+, the Rb+ ion is bound with unfavorable geometry, and it is proposed that the rigid site 2 structure results in a pronounced selectivity for Na+ ions.
磷酸吡哆醛依赖性酶二烷基甘氨酸脱羧酶（DGD）被K⁺和Rb⁺离子激活，而Li⁺和Na⁺离子则具有抑制作用。在DGD的晶体结构中发现了一个靠近活性位点（位点1）的碱金属离子结合位点，并且该位点上K⁺与Na⁺的交换被证明会影响两个活性位点残基的构象 [托尼，M. D.，霍内斯特，E.，考恩，S. W.，& 扬松纽斯，J. N.（1993年）《科学》261卷，756 - 759页]。我们研究了碱金属离子对DGD活性的影响，并确定了位点1结合Li⁺和Rb⁺的DGD在2.8 Å分辨率下的晶体结构。由于Li⁺离子的散射较弱，其位置必须使用小分子结构的信息进行建模。对位点1结合Li⁺、Na⁺、K⁺和Rb⁺的DGD结构进行比较，揭示了活性位点结构与酶活性之间的显著相关性。较小的抑制性离子Li⁺和Na⁺通过单个水分子取代较大的激活离子K⁺和Rb⁺的两个蛋白质衍生配体来容纳。这引发了活性酶和非活性酶之间的双态结构转换，涉及活性位点残基Ser80和Tyr301的协同重新定向以及DGD四聚体四级结构的微小变化。提出了必需的K⁺离子在辅因子结合和催化活性位点结构组织中的重要作用。在位点
1结合Rb⁺的DGD结构中，第二个Rb⁺离子部分取代了DGD分子表面金属结合位点2上的结构Na⁺离子，而没有显著改变蛋白质结构。与Na⁺不同，Rb⁺离子以不利的几何构型结合，并且提出刚性的位点2结构导致对Na⁺离子有明显的选择性。
