Sakuraba Haruhiko, Tsuge Hideaki, Yoneda Kazunari, Katunuma Nobuhiko, Ohshima Toshihisa
Department of Biological Science and Technology, Faculty of Engineering, The University of Tokushima, 2-1 Minamijosanjima-cho, Tokushima 770-8506.
J Biol Chem. 2005 Jul 22;280(29):26645-8. doi: 10.1074/jbc.C500192200. Epub 2005 Jun 3.
A gene encoding a quinolinate synthase has been identified in the hyperthermophilic archaeon Pyrococcus horikoshii via genome sequencing. The gene was overexpressed in Escherichia coli, and the crystal structure of the produced enzyme was determined to 2.0 A resolution in the presence of malate, a substrate analogue. The overall structure exhibits a unique triangular architecture composed of a 3-fold repeat of three-layer (alphabetaalpha) sandwich folding. Although some aspects of the fold homologous to the each domain have been observed previously, the overall structure of quinolinate synthase shows no similarity to any known protein structure. The three analogous domains are related to a pseudo-3-fold symmetry. The active site is located at the interface of the three domains and is centered on the pseudo-3-fold axis. The malate molecule is tightly held near the bottom of the active site cavity. The model of the catalytic state during the first condensation step of the quinolinate synthase reaction indicates that the elimination of inorganic phosphate from dihydroxyacetone phosphate may precede the condensation reaction.
通过基因组测序，在嗜热古菌火之神 Pyrococcus horikoshii 中鉴定出了一种编码喹啉酸合酶的基因。该基因在大肠杆菌中过表达，并在底物类似物苹果酸存在的情况下，将所产生酶的晶体结构解析到 2.0 Å 的分辨率。整体结构呈现出一种独特的三角形架构，由三层（αβ α）三明治折叠的三倍重复组成。尽管先前已经观察到与每个结构域同源的折叠的某些方面，但喹啉酸合酶的整体结构与任何已知蛋白质结构均无相似性。三个类似的结构域与假三重对称相关。活性位点位于三个结构域的界面处，并以假三重轴为中心。苹果酸分子紧密地保持在活性位点腔底部附近。喹啉酸合酶反应第一步缩合过程中的催化状态模型表明，从磷酸二羟丙酮中消除无机磷酸可能先于缩合反应。
