Sequential events in the irreversible thermal denaturation of human brain-type creatine kinase by spectroscopic methods.
Sequential events in the irreversible thermal denaturation of human brain-type creatine kinase by spectroscopic methods.
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用光谱方法研究人脑型肌酸激酶不可逆热变性的序列事件
DOI：
10.3390/ijms11072584
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发表时间：
2010-06-25
影响因子：
5.6
通讯作者：
Yan YB
中科院分区：
文献类型：
--
作者：
Gao YS;Su JT;Yan YB
蛋白质热变性过程中发生的非合作或连续事件与蛋白质折叠、稳定性和生理功能密切相关。在本研究中，通过差示扫描量热法 (DSC)、CD 和内在荧光光谱研究了人脑型肌酸激酶 (hBBCK) 热变性的连续事件。 DSC 实验表明 hBBCK 的热变性是量热不可逆的。多个吸热峰的存在表明变性涉及逐步构象变化，这通过从各种光谱探针获得的转变曲线的差异得到进一步验证。在加热过程中，活性位点结构的破坏发生在二级和三级结构变化之前。发现 hBBCK 的热解折叠和聚集是通过连续事件发生的。这与肌肉型CK（MMCK）有很大不同。本文的结果表明，BBCK 和 MMCK 经历非常不同的热解折叠途径，尽管它们在一级和三级结构中高度保守。结构上的微小差异可能赋予同工酶不同的局部结构稳定性，这进一步有助于同工酶特异性的热稳定性。
The non-cooperative or sequential events which occur during protein thermal denaturation are closely correlated with protein folding, stability, and physiological functions. In this research, the sequential events of human brain-type creatine kinase (hBBCK) thermal denaturation were studied by differential scanning calorimetry (DSC), CD, and intrinsic fluorescence spectroscopy. DSC experiments revealed that the thermal denaturation of hBBCK was calorimetrically irreversible. The existence of several endothermic peaks suggested that the denaturation involved stepwise conformational changes, which were further verified by the discrepancy in the transition curves obtained from various spectroscopic probes. During heating, the disruption of the active site structure occurred prior to the secondary and tertiary structural changes. The thermal unfolding and aggregation of hBBCK was found to occur through sequential events. This is quite different from that of muscle-type CK (MMCK). The results herein suggest that BBCK and MMCK undergo quite dissimilar thermal unfolding pathways, although they are highly conserved in the primary and tertiary structures. A minor difference in structure might endow the isoenzymes dissimilar local stabilities in structure, which further contribute to isoenzyme-specific thermal stabilities.
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