Watson K A, Schinzel R, Palm D, Johnson L N
Laboratory of Molecular Biophysics, Oxford, UK.
EMBO J. 1997 Jan 2;16(1):1-14. doi: 10.1093/emboj/16.1.1.
In animals, glycogen phosphorylase (GP) exists in an inactive (T state) and an active (R state) equilibrium that can be altered by allosteric effectors or covalent modification. In Escherichia coli, the activity of maltodextrin phosphorylase (MalP) is controlled by induction at the level of gene expression, and the enzyme exhibits no regulatory properties. We report the crystal structure of E. coli maltodextrin phosphorylase refined to 2.4 A resolution. The molecule consists of a dimer with 796 amino acids per monomer, with 46% sequence identity to the mammalian enzyme. The overall structure of MalP shows a similar fold to GP and the catalytic sites are highly conserved. However, the relative orientation of the two subunits in E. coli MalP is different from both the T and R state GP structures, and there are significant changes at the subunit-subunit interfaces. The sequence changes result in loss of each of the control sites present in rabbit muscle GP. As a result of the changes at the subunit interface, the 280s loop, which in T state GP acts as a gate to control access to the catalytic site, is held in an open conformation in MalP. The open access to the conserved catalytic site provides an explanation for the activity without control in this basic archetype of a phosphorylase.
在动物体内，糖原磷酸化酶（GP）以无活性（T态）和活性（R态）的平衡状态存在，这种平衡可通过变构效应物或共价修饰来改变。在大肠杆菌中，麦芽糖糊精磷酸化酶（MalP）的活性受基因表达水平的诱导控制，且该酶不具有调节特性。我们报道了分辨率为2.4埃的大肠杆菌麦芽糖糊精磷酸化酶的晶体结构。该分子由二聚体组成，每个单体含有796个氨基酸，与哺乳动物的酶具有46%的序列同一性。MalP的整体结构与GP具有相似的折叠方式，且催化位点高度保守。然而，大肠杆菌MalP中两个亚基的相对取向与T态和R态的GP结构均不同，并且在亚基-亚基界面处存在显著变化。序列变化导致兔肌肉GP中存在的每个控制位点丧失。由于亚基界面处的变化，在T态GP中作为控制进入催化位点之门的280s环在MalP中保持开放构象。对保守催化位点的开放访问为这种磷酸化酶基本原型中不受控制的活性提供了解释。
