胞苷-5'-三磷酸(CTP)合酶(CTPS)是I类谷氨酰胺依赖性酰胺基转移酶(GAT),催化CTP从头生物合成的最后一步。谷氨酰胺水解在GAT结构域中被催化,并且释放的氨经由分子内隧道转移至合酶结构域,其中发生UTP的ATP依赖性胺化以形成CTP。CTPS在谷氨酰胺依赖性酰胺转移酶中是独一无二的,需要变构效应子(GTP)来激活GAT结构域以进行有效的谷氨酰胺水解。最近,果蝇CTPS的第一个低温电子显微镜结构是用结合的ATP解决的,UTP,and,特别是,GTP,以及与6-重氮-5-氧代-1-正亮氨酸的共价加合物。这些结构信息,随着大量的定点诱变,动力学,以及过去50年进行的结构研究,提供有关GTP在GAT域结合的复杂构象变化及其对催化的贡献的更详细见解。GTP和L2环路之间的交互,来自相邻质子器的L4循环,L11盖子,和L13环路(或独特的柔性“机翼”区域),诱导构象变化,促进谷氨酰胺在GAT结构域的水解;然而,仍然缺乏有关这些构象变化促进GAT结构域催化的特定机制的直接实验证据。重要的是,GTP结合诱导的构象变化也影响NH3隧道的组装和维持。因此,除了促进谷氨酰胺水解,变构效应子在协调由CTPS的GAT和合酶结构域催化的反应中起着重要作用。
Cytidine-5\'-triphosphate (CTP) synthase (CTPS) is the class I glutamine-dependent amidotransferase (GAT) that catalyzes the last step in the de novo biosynthesis of CTP. Glutamine hydrolysis is catalyzed in the GAT domain and the liberated ammonia is transferred via an intramolecular tunnel to the synthase domain where the ATP-dependent amination of UTP occurs to form CTP. CTPS is unique among the glutamine-dependent amidotransferases, requiring an allosteric effector (GTP) to activate the GAT domain for efficient glutamine hydrolysis. Recently, the first cryo-electron microscopy structure of Drosophila CTPS was solved with bound ATP, UTP, and, notably, GTP, as well as the covalent adduct with 6-diazo-5-oxo-l-norleucine. This structural information, along with the numerous site-directed mutagenesis, kinetics, and structural studies conducted over the past 50 years, provide more detailed insights into the elaborate conformational changes that accompany GTP binding at the GAT domain and their contribution to catalysis. Interactions between GTP and the L2 loop, the L4 loop from an adjacent protomer, the L11 lid, and the L13 loop (or unique flexible \"wing\" region), induce conformational changes that promote the hydrolysis of glutamine at the GAT domain; however, direct experimental evidence on the specific mechanism by which these conformational changes facilitate catalysis at the GAT domain is still lacking. Significantly, the conformational changes induced by GTP binding also affect the assembly and maintenance of the NH3 tunnel. Hence, in addition to promoting glutamine hydrolysis, the allosteric effector plays an important role in coordinating the reactions catalyzed by the GAT and synthase domains of CTPS.