Dissecting the pretransitional conformational changes in aminoacylase I thermal denaturation
Dissecting the pretransitional conformational changes in aminoacylase I thermal denaturation
复制标题
剖析氨酰化酶 I 热变性中的过渡前构象变化
DOI：
10.1529/biophysj.106.093666
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发表时间：
2007-01-01
影响因子：
3.4
通讯作者：
Yan, Yong-Bin
中科院分区：
文献类型：
--
作者：
Su, Jing-Tan;Kim, Sung-Hye;Yan, Yong-Bin
氨基酰化酶 I (ACYI) 催化 L-酰基氨基酸的立体特异性水解，并且通常被认为参与细胞内 N-乙酰化蛋白质降解的最后步骤。除了在细胞内氨基酸代谢中的关键功能外，ACYI 对于 N-酰化 DL-氨基酸的光学拆分也具有重大的商业重要性。作为一种锌依赖性酶，ACYI 对热变性非常稳定，可以被视为一种热稳定酶，其最佳活性温度约为 65°C。在本研究中，通过结合光谱方法和相关分辨率增强技术研究了 ACYI 热变性的连续事件。有趣的是，结果来自。荧光和红外（IR）光谱清楚地表明，在50℃至66℃的温度下存在过渡前阶段。ACYI的热解折叠可能是一个三态过程，涉及在大约68℃下出现的易于聚集的中间体。过渡前结构变化涉及溶剂暴露的β-折叠结构的部分解折叠以及大约一半的I类Trp荧光团向II类的转变。我们的结果还表明，使用分辨率增强技术可以提供蛋白质逐步展开的有价值的信息。
Aminoacylase I (ACYI) catalyzes the stereospecific hydrolysis of L-acylamino acids and is generally assumed to be involved in the final step of the degradation of intracellular N-acetylated proteins. Apart from its crucial functions in intracellular amino acid metabolism, ACYI also has substantial commercial importance for the optical resolution of N-acylated DL-amino acids. As a zinc-dependent enzyme, ACYI is quite stable against heat-induced denaturation and can be regarded as a thermostable enzyme with an optimal temperature for activity of similar to 65 degrees C. In this research, the sequential events in ACYI thermal denaturation were investigated by a combination of spectroscopic methods and related resolution-enhancing techniques. Interestingly, the results from. uorescence and infrared (IR) spectroscopy clearly indicated that a pretransitional stage existed at temperatures from 50 degrees C to 66 degrees C. The thermal unfolding of ACYI might be a three-state process involving an aggregation-prone intermediate appearing at similar to 68 degrees C. The pretransitional structural changes involved the partial unfolding of the solvent-exposed beta-sheet structures and the transformation of about half of the Class I Trp fluorophores to Class II. Our results also suggested that the usage of resolution-enhancing techniques could provide valuable information of the step-wise unfolding of proteins.