REFINED CRYSTAL-STRUCTURE OF LIPOAMIDE DEHYDROGENASE FROM AZOTOBACTER-VINELANDII AT 2.2-A RESOLUTION - A COMPARISON WITH THE STRUCTURE OF GLUTATHIONE-REDUCTASE
REFINED CRYSTAL-STRUCTURE OF LIPOAMIDE DEHYDROGENASE FROM AZOTOBACTER-VINELANDII AT 2.2-A RESOLUTION - A COMPARISON WITH THE STRUCTURE OF GLUTATHIONE-REDUCTASE
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DOI：
10.1016/0022-2836(91)90367-f
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发表时间：
1991-08-20
影响因子：
5.6
通讯作者：
HOL, WGJ
中科院分区：
文献类型：
--
作者：
MATTEVI, A;SCHIERBEEK, AJ;HOL, WGJ
用分子动力学方法对棕色固氮菌硫辛酰胺脱氢酶的结构进行了精细化，在2.2 μ m分辨率下，R因子为19.8%。在最终的模型中，键长和键角的均方根偏差分别为0.02 °和3.2 °。不对称单元包括两个亚基，每个亚基由466个氨基酸残基和辅基FAD加上512个溶剂分子组成。两条链的最后10个氨基酸残基在电子密度分布中不可见，它们可能是无序的。使形成二聚体的两条链相互作用所需的操作是精确地旋转180 °，而没有平移分量。最后的模型表明，两个独立的细化亚基是非常相似的，除了位于分子表面的六个环。每个亚基的酶的结构由四个结构域与催化中心位于亚基界面。反应性二硫桥48-53被Cys 53的Sγ氧化，该Sγ位于异咯嗪环的碳C-4a的3.5 nm处。His 450 ′侧链的Nε 2指向Cys 48的Sγ，并与Glu 455 ′的羧基形成氢键。FAD以伸展构象结合，异咯嗪环不是完全平面的，蝶啶与苯环的夹角在第一亚基为7.3 °，在第二亚基为12.1 °，硫辛酰胺脱氢酶的整体折叠与谷胱甘肽还原酶的折叠非常相似。然而，这两种酶，其中只有26%的序列同一性的比较，揭示了显着的构象差异。这些关系到这两个分子的三级和四级结构。在硫辛酰胺脱氢酶的每个亚基中，NAD结合结构域和界面结构域相对于FAD结合结构域的取向分别不同7.1 °和7.8 °。此外，界面域还包含三级结构的主要变化。此外，硫辛酰胺脱氢酶二聚体与谷胱甘肽还原酶二聚体相比，形成二聚体的两个亚基似乎相对于彼此移位超过4 μ m。尽管所有这些变化在三级和四级水平的酶的活性位点，发生在二聚体界面，似乎是非常相似的。显然，观察到的许多构象变化以这样一种方式相互补偿，即活性位点中的关键残基在这两种酶中具有相同的相互位置和方向。
The structure of lipoamide dehydrogenase fromAzotobacter vinelandiihas been refined by the molecular dynamics technique to anR-factor of 19.8% at 2.2 Å resolution. In the final model, the root-mean-square deviation from ideality is 0.02 Å for bond lengths and 3.2 ° for bond angles. The asymmetric unit comprises two subunits, each consisting of 466 amino acid residues and the prosthetic group FAD, plus 512 solvent molecules. The last ten amino acid residues of both chains are not visible in the electron density distribution and they are probably disordered. The operation required to superimpose the two chains forming the dimer is a rotation of exactly 180 ° with no translation component. The final model shows the two independently refined subunits to be very similar, except for six loops located at the surface of the molecule.The structure of each subunit of the enzyme consists of four domains with the catalytic centre located at the subunit interface. The reactive disulphide bridge, 48–53, is oxidized with Sγof Cys53 located 3.5 Å away from carbon C-4a of the isoalloxazine ring. The side-chain of His450′ points its Nε2towards Sγof Cys48 and is hydrogen bonded to the carboxylate of Glu455′. The FAD is bound in an extended conformation and the isoalloxazine ring is not completely planar with an angle between the pteridine and the benzene ring of 7.3 ° in the first subunit and of 12.1 ° in the second one.The overall folding of lipoamide dehydrogenase is very similar to that of glutathione reductase. However, a comparison of the two enzymes, which have only 26% sequence identity, reveals significant conformational differences. These concern the tertiary as well as the quaternary structure of the two molecules. In each subunit of lipoamide dehydrogenase the NAD-binding domain and the interface domain appear to be differently oriented with respect to the FAD-binding domain by 7.1 ° and 7.8 °, respectively. The interface domain contains, in addition, major changes in tertiary structure. Furthermore, the two subunits forming the dimer appear to be shifted with respect to each other by more than 4 Å, when the lipoamide dehydrogenase dimer is compared with that of glutathione reductase. In spite of all these changes at the tertiary and quaternary level the active sites of the enzymes, which occur at the dimer interface, appear to be remarkably similar. Clearly, many of the conformational changes observed compensate each other in such a manner that key residues in the active site have the same mutual position and orientation in these two enzymes.