Scapin G, Cirilli M, Reddy S G, Gao Y, Vederas J C, Blanchard J S
Biochemistry Department, Albert Einstein College of Medicine, New York 10469, USA.
Biochemistry. 1998 Mar 10;37(10):3278-85. doi: 10.1021/bi9727949.
The three-dimensional structures of Corynebacterium glutamicum diaminopimelate dehydrogenase as a binary complex with the substrate meso-diaminopimelate (meso-DAP) and a ternary complex with NADP+ and an isoxazoline inhibitor [Abbot, S.D., Lane-Bell, P., Kanwar, P.S.S., and Vederas, J. C. (1994) J. Am. Chem. Soc. 116, 6513-6520] have been solved and refined against X-ray diffraction data to 2.2 A. Diaminopimelate dehydrogenase is a homodimer of approximately 35,000 molecular weight subunits and is the only dehydrogenase present in the bacterial diaminopimelate/lysine biosynthetic pathway. Inhibitors of the enzymes of L-lysine biosynthesis have been proposed as potential antibiotics or herbicides, since mammals lack this metabolic pathway. Diaminopimelate dehydrogenase catalyzes the unique, reversible, pyridine dinucleotide-dependent oxidative deamination of the D-amino acid stereocenter of meso-diaminopimelate to generate L-2-amino-6-oxopimelate. The enzyme is absolutely specific for the meso stereoisomer of DAP and must distinguish between two opposite chiral amino acid centers on the same symmetric substrate. The determination of the three-dimensional structure of the enzyme--meso-diaminopimelate complex allows a description of the molecular basis of this stereospecific discrimination. The substrate is bound in an elongated cavity, in which the distribution of residues that act as hydrogen bond donors or acceptors defines a single orientation in which the substrate may bind in order to position the D-amino acid center of meso-DAP near the oxidized nucleotide. The previously described isoxazoline inhibitor binds at the same site as DAP but has its L-amino acid center positioned where the D-amino acid center of meso-DAP would normally be located, thereby generating a nonproductive inhibitor complex. The relative positions of the N-terminal dinucleotide and C-terminal substrate-binding domains in the diaminopimelate dehydrogenase--NADP+, diaminopimelate dehydrogenase--DAP, and diaminopimelate dehydrogenase--NADP(+)--inhibitor complexes confirm our previous observations that the enzyme undergoes significant conformational changes upon binding of both dinucleotide and substrate.
谷氨酸棒杆菌二氨基庚二酸脱氢酶与底物内消旋二氨基庚二酸（meso-DAP）形成的二元复合物以及与NADP⁺和异恶唑啉抑制剂形成的三元复合物的三维结构已被解析，并根据X射线衍射数据精修至2.2 Å。二氨基庚二酸脱氢酶是由分子量约为35,000的亚基组成的同型二聚体，是细菌二氨基庚二酸/赖氨酸生物合成途径中唯一存在的脱氢酶。由于哺乳动物缺乏这种代谢途径，L-赖氨酸生物合成酶的抑制剂已被提议作为潜在的抗生素或除草剂。二氨基庚二酸脱氢酶催化内消旋二氨基庚二酸的D-氨基酸立体中心独特的、可逆的、依赖吡啶二核苷酸的氧化脱氨反应，生成L-2-氨基-6-氧代庚二酸。该酶对DAP的内消旋立体异构体具有绝对特异性，并且必须区分同一对称底物上的两个相反手性氨基酸中心。酶 - 内消旋二氨基庚二酸复合物三维结构的确定使得能够描述这种立体特异性识别的分子基础。底物结合在一个细长的腔内，其中作为氢键供体或受体的残基分布定义了底物可能结合的单一取向，以便将meso-DAP的D-氨基酸中心定位在氧化核苷酸附近。先前描述的异恶唑啉抑制剂与DAP结合在同一位置，但它的L-氨基酸中心位于meso-DAP的D-氨基酸中心通常所在的位置，从而形成非生产性抑制剂复合物。二氨基庚二酸脱氢酶 - NADP⁺、二氨基庚二酸脱氢酶 - DAP和二氨基庚二酸脱氢酶 - NADP⁺ - 抑制剂复合物中N端二核苷酸和C端底物结合结构域的相对位置证实了我们之前的观察结果，即该酶在结合二核苷酸和底物时会发生显著的构象变化。
