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首页> 外文期刊>Journal of Molecular Biology >Structural and functional properties of isocitrate dehydrogenase from the psychrophilic bacterium Desulfotalea psychrophila reveal a cold-active enzyme with an unusual high thermal stability
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Structural and functional properties of isocitrate dehydrogenase from the psychrophilic bacterium Desulfotalea psychrophila reveal a cold-active enzyme with an unusual high thermal stability

机译：嗜冷细菌脱硫嗜水杆菌异柠檬酸脱氢酶的结构和功能特性揭示了一种具有异常高热稳定性的冷活性酶

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Isocitrate dehydrogenase (IDH) has been studied extensively due to its central role in the Krebs cycle, catalyzing the oxidative NAD(P)(+)-dependent decarboxylation of isocitrate to alpha-ketoglutarate and CO2. Here, we present the first crystal structure of IDH from a psychrophilic bacterium, Desulfotalea psychrophila (DpIDH). The structural information is combined with a detailed biochemical characterization and a comparative study with IDHs (PcIDH), human cytosolic (HcIDH) and the hyperthermophilic Thermotoga maritima (TmIDH). DpIDH was found to have a higher melting temperature (T-m = 66.9 degrees C) than its mesophilic homologues and a suboptimal catalytic efficiency at low temperatures. The thermodynamic activation parameters indicated a disordered active site, as seen also for the drastic increase in Km for isocitrate at elevated temperatures. A methionine cluster situated at the dimeric interface between the two active sites and a cluster of destabilizing charged amino acids in a region close to the active site might explain the poor isocitrate affinity. On the other hand, DpIDH was optimized for interacting with NADP(+) and the crystal structure revealed unique interactions with the cofactor. The highly acidic surface, destabilizing charged residues, fewer ion pairs and reduced size of ionic networks in DpIDH suggest a flexible global structure. However, strategic placement of ionic interactions stabilizing the N and C termini, and additional ionic interactions in the clasp domain as well as two enlarged aromatic clusters might counteract the destabilizing interactions and promote the increased thermal stability. The structure analysis of DpIDH illustrates how psychrophilic enzymes can adjust their flexibility in dynamic regions during their catalytic cycle without compromising the global stability of the protein. (c) 2007 Elsevier Ltd. All rights reserved.
机译：异柠檬酸脱氢酶（IDH）由于其在克雷布斯循环中的核心作用而被广泛研究，该反应催化异柠檬酸的氧化NAD（P）（+）依赖性脱羧反应生成α-酮戊二酸和CO2。在这里，我们介绍了来自嗜冷细菌，脱硫嗜水菌（DsIDotaH Psychrophila，DpIDH）的IDH的第一个晶体结构。该结构信息与详细的生化特征以及与IDH（PcIDH），人胞质（HcIDH）和超嗜热嗜热栖热球菌（TmIDH）的比较研究相结合。发现DpIDH具有比其嗜温同源物更高的熔化温度（T-m = 66.9摄氏度），并且在低温下催化效果不佳。热力学活化参数表明活性位点无序，如在升高的温度下异柠檬酸钾的Km急剧增加也可见。位于两个活性位点之间的二聚体界面的甲硫氨酸簇和靠近活性位点的区域中不稳定电荷的氨基酸簇可能解释了异柠檬酸亲和力差。另一方面，DpIDH被优化为与NADP（+）相互作用，并且晶体结构显示与辅因子的独特相互作用。 DpIDH中高度酸性的表面，不稳定的带电残基，更少的离子对和离子网络的尺寸减小，提示了灵活的全局结构。但是，稳定N和C末端的离子相互作用的策略性位置以及扣环域以及两个扩大的芳族簇中的其他离子相互作用可能会抵消不稳定的相互作用，并提高热稳定性。 DpIDH的结构分析表明，嗜冷酶如何在其催化循环过程中调节其在动态区域中的柔韧性，而不会损害该蛋白的整体稳定性。 （c）2007 Elsevier Ltd.保留所有权利。

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