Závodszky P, Kardos J, Petsko G A
Institute of Enzymology, Biological Research Center, Hungarian Academy of Sciences, Pf. 7, H-1518 Budapest, Hungary.
Proc Natl Acad Sci U S A. 1998 Jun 23;95(13):7406-11. doi: 10.1073/pnas.95.13.7406.
3-Isopropylmalate dehydrogenase (IPMDH, E.C. 1.1.1.85) from the thermophilic bacterium Thermus thermophilus HB8 is homologous to IPMDH from the mesophilic Escherichia coli, but has an approximately 17 degreesC higher melting temperature. Its temperature optimum is 22-25 degreesC higher than that of the E. coli enzyme; however, it is hardly active at room temperature. The increased conformational rigidity required to stabilize the thermophilic enzyme against heat denaturation might explain its different temperature-activity profile. Hydrogen/deuterium exchange studies were performed on this thermophilic-mesophilic enzyme pair to compare their conformational flexibilities. It was found that Th. thermophilus IPMDH is significantly more rigid at room temperature than E. coli IPMDH, whereas the enzymes have nearly identical flexibilities under their respective optimal working conditions, suggesting that evolutionary adaptation tends to maintain a "corresponding state" regarding conformational flexibility. These observations confirm that conformational fluctuations necessary for catalytic function are restricted at room temperature in the thermophilic enzyme, suggesting a close relationship between conformational flexibility and enzyme function.
嗜热栖热菌HB8的3-异丙基苹果酸脱氢酶（IPMDH，酶委员会编号1.1.1.85）与嗜温性大肠杆菌的IPMDH同源，但其解链温度高出约17摄氏度。其最适温度比大肠杆菌的该酶高22 - 25摄氏度；然而，它在室温下几乎没有活性。稳定嗜热酶以抵抗热变性所需的构象刚性增加可能解释了其不同的温度 - 活性曲线。对这对嗜热 - 嗜温酶进行了氢/氘交换研究，以比较它们的构象灵活性。结果发现，嗜热栖热菌的IPMDH在室温下比大肠杆菌的IPMDH刚性显著更高，而这两种酶在各自的最佳工作条件下具有几乎相同的灵活性，这表明进化适应倾向于在构象灵活性方面维持一种“对应状态”。这些观察结果证实，嗜热酶在室温下催化功能所需的构象波动受到限制，这表明构象灵活性与酶功能之间存在密切关系。
