Solution structure of the phosphoryl transfer complex between the signal-transducing protein IIAGlucose and the cytoplasmic domain of the glucose transporter IICBGlucose of the Escherichia coli glucose phosphotransferase system
Solution structure of the phosphoryl transfer complex between the signal-transducing protein IIAGlucose and the cytoplasmic domain of the glucose transporter IICBGlucose of the Escherichia coli glucose phosphotransferase system
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DOI：
10.1074/jbc.m302677200
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发表时间：
2003-07-04
影响因子：
4.8
通讯作者：
Clore, GM
中科院分区：
文献类型：
--
作者：
Cai, ML;Williams, DC;Clore, GM
在大肠杆菌磷酸转移酶系统的葡萄糖特异性臂中，在酶IIA（葡萄糖）（IIA（Glc））和葡萄糖转运蛋白IICBGlc的细胞质B结构域（IIBGlc）之间的最终磷酰基转移复合物的溶液结构已经通过NMR解析。该界面（类似于1200埃（2）埋面）是由IIA（Glc）上的凹陷与IIBGlc上的凸起相互作用形成的。磷酰基供体和受体残基，IIA（Glc）的His-90和IIBGlc的Cys-35（IIBGlc的残基以斜体表示）非常接近并埋在界面的中心。Cys-35通过与几个相邻骨架酰胺基团的分子内氢键相互作用稳定硫醇盐阴离子（pK（a）类似于6.5），从而引发对磷原子的亲核攻击。疏水性分子间接触的补充外周静电相互作用，涉及交替分布的正电荷和负电荷的残基的相互作用表面上的两种蛋白质。IIA（Glc）的Asp-38/Asp-94对和IIBGlc的Arg-38/Arg-40对之间的盐桥中和了复合物中Cys-35的S γ原子和磷酰基附近的负电荷积累。一个五配位磷酰基过渡态是很容易容纳的骨架构象没有任何变化，和复杂的结构帐户之间的IIA（葡萄糖）和IIBGlc的磷酰基转移的首选方向性。IIA（Glc）.IIBGlc和葡萄糖磷酸转移酶系统的两个上游复合物（EI.HPr和IIA（Glc）.HPr）的结构揭示了其中HPr和IIA（Glc）上高度重叠的结合位点识别结构不同的蛋白质的级联。
The solution structure of the final phosphoryl transfer complex in the glucose-specific arm of the Escherichia coli phosphotransferase system, between enzyme IIA(Glucose) (IIA(Glc)) and the cytoplasmic B domain (IIBGlc) of the glucose transporter IICBGlc, has been solved by NMR. The interface (similar to1200-Angstrom(2) buried surface) is formed by the interaction of a concave depression on IIA(Glc) with a convex protrusion on IIBGlc. The phosphoryl donor and acceptor residues, His-90 of IIA(Glc) and Cys-35 of IIBGlc ( residues of IIBGlc are denoted in italics) are in close proximity and buried at the center of the interface. Cys-35 is primed for nucleophilic attack on the phosphorus atom by stabilization of the thiolate anion (pK(a) similar to6.5) through intramolecular hydrogen bonding interactions with several adjacent backbone amide groups. Hydrophobic intermolecular contacts are supplemented by peripheral electrostatic interactions involving an alternating distribution of positively and negatively charged residues on the interaction surfaces of both proteins. Salt bridges between the Asp-38/Asp-94 pair of IIA(Glc) and the Arg-38/Arg-40 pair of IIBGlc neutralize the accumulation of negative charge in the vicinity of both the Sgamma atom of Cys-35 and the phosphoryl group in the complex. A pentacoordinate phosphoryl transition state is readily accommodated without any change in backbone conformation, and the structure of the complex accounts for the preferred directionality of phosphoryl transfer between IIA(Glc) and IIBGlc. The structures of IIA(Glc).IIBGlc and the two upstream complexes of the glucose phosphotransferase system (EI.HPr and IIA(Glc).HPr) reveal a cascade in which highly overlapping binding sites on HPr and IIA(Glc) recognize structurally diverse proteins.