Magyar C, Szilágyi A, Závodszky P
Institute of Enzymology, Biological Research Center, Hungarian Academy of Sciences, Budapest, Hungary.
Protein Eng. 1996 Aug;9(8):663-70. doi: 10.1093/protein/9.8.663.
To reveal the structural basis of the increased thermal stability of 3-isopropylmalate dehydrogenase (IPMDH) from Thermus thermophilus, an extreme thermophile, the homology-based structural model of one mesophilic (Escherichia coli) counterpart, was constructed. Both IPMDHs are homodimeric proteins. We built a model of one subunit using the 3-D structures of the Th. thermophilus IPMDH and the homologous E.coli isocitrate dehydrogenase. Energy minimization and molecular dynamics simulated annealing were performed on the dimer, including a surrounding solvation shell. No serious errors were detected in the refined model using the 3-D profile method. The resulting structure was scrutinized and compared with the structure of the Th.thermophilus IPMDH. Significant differences were found in the non-specific interactions including the hydrophobic effect. The model predicts a higher number of ion pairs in the Th.thermophilus than in the E.coli enzyme. An increase was observed in the stabilities of alpha-helical regions in the thermophilic protein. The preliminary X-ray coordinates of the E.coli IPMDH were received after the completion of this work, allowing an assessment of the model in terms of the X-ray structure. The comparison proved that most of the structural features underlying the stability differences between the two enzymes were predicted correctly.
为揭示嗜热栖热菌（一种极端嗜热菌）中3-异丙基苹果酸脱氢酶（IPMDH）热稳定性增加的结构基础，构建了一种嗜温菌（大肠杆菌）对应物基于同源性的结构模型。两种IPMDH均为同二聚体蛋白。我们利用嗜热栖热菌IPMDH和同源的大肠杆菌异柠檬酸脱氢酶的三维结构构建了一个亚基的模型。对包括周围溶剂化层在内的二聚体进行了能量最小化和分子动力学模拟退火。使用三维轮廓法在优化后的模型中未检测到严重错误。对所得结构进行了仔细检查，并与嗜热栖热菌IPMDH的结构进行了比较。在包括疏水作用在内的非特异性相互作用中发现了显著差异。该模型预测嗜热栖热菌中的离子对比大肠杆菌酶中的更多。在嗜热蛋白的α-螺旋区域的稳定性中观察到增加。在这项工作完成后收到了大肠杆菌IPMDH的初步X射线坐标，从而能够根据X射线结构对模型进行评估。比较证明，两种酶之间稳定性差异的大多数结构特征都被正确预测。
