Del Vecchio Pompea, Graziano Giuseppe, Barone Guido
Dipartimento di Chimica, Università di Napoli Federico II, Via Cintia, 80126 Napoli, Italy.
Protein Pept Lett. 2009;16(10):1201-6. doi: 10.2174/092986609789071315.
In the last years we have performed a series of studies to characterize the conformational stability of three esterases from thermophilic and mesophilic sources: Aes esterase from Escherichia coli, EST2 from Alicyclobacillus acidocaldarius and AFEST from Archeoglobus fulgidus. These three esterases belong to the Hormone-sensitive lipase group of the superfamily of carboxylester hydrolases. The conformational stability of the three enzymes against temperature, urea and GuHCl has been determined by means of circular dichroism, fluorescence and differential scanning calorimetry measurements. Analysis of experimental data coupled with available structural information allowed us to suggest that the optimization of charge-charge interactions on the protein surface could one of the mechanisms to increase the thermal stability for the three esterases. This idea has been tested in the case of EST2, which shows a fully reversible thermal unfolding, by producing and studying variant forms of wild type enzyme in which a charged residue has been mutated. In the present article the obtained results are critically recollected in order to provide a clear and unified scenario.
在过去几年中，我们开展了一系列研究，以表征来自嗜热和嗜温源的三种酯酶的构象稳定性：大肠杆菌的Aes酯酶、嗜酸嗜热栖热菌的EST2以及嗜热栖热菌的AFEST。这三种酯酶属于羧酸酯水解酶超家族的激素敏感脂肪酶组。通过圆二色性、荧光和差示扫描量热法测量，确定了这三种酶对温度、尿素和盐酸胍的构象稳定性。对实验数据的分析以及可用的结构信息使我们能够提出，优化蛋白质表面的电荷-电荷相互作用可能是提高这三种酯酶热稳定性的机制之一。在EST2的情况下，这一想法已通过产生并研究野生型酶的变体形式（其中一个带电荷的残基发生了突变）进行了验证，EST2表现出完全可逆的热解折叠。在本文中，对获得的结果进行了批判性回顾，以提供一个清晰统一的情况。
