The molecular basis of the effect of temperature on enzyme activity
The molecular basis of the effect of temperature on enzyme activity
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DOI：
10.1042/bj20091254
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发表时间：
2010-01-15
影响因子：
4.1
通讯作者：
Lee, Charles K.
中科院分区：
文献类型：
--
作者：
Daniel, Roy M.;Peterson, Michelle E.;Lee, Charles K.
实验数据表明，温度对酶的影响不能用基于活性和变性增加的二态模型来充分解释。平衡模型通过引入与活性形式快速平衡的非活性(但未变性)中间体，提供了反应条件下酶热行为的定量解释。这两种形式之间快速平衡的温度中点(T-eq)与生物体的生长温度有关，平衡的热焓(Delta H-eq)与其在不同温度范围内发挥作用的能力有关。在本研究中，我们证明了活性形式和非活性形式之间的差异在于酶的活性部位。结果揭示了一种明显的普遍机制，与酶的反应或结构无关，基于活性部位或附近，酶通过该机制随着温度的升高而失去活性，而不是全球变性。结果表明，T-eq以下的活性损失可能会导致根据两态(经典)模型测定Delta G(CAT)*的显著误差，所测得的k(CAT)将不能真实地指示酶的催化能力。总体而言，这些结果为观察到活性位置往往比酶整体更灵活提供了分子基础，并且活性损失先于变性，并从分子角度对温度对酶活性的影响提供了一般解释。
Experimental data show that the effect of temperature on enzymes cannot be adequately explained in terms of a two-state model based on increases in activity and denaturation. The Equilibrium Model provides a quantitative explanation of enzyme thermal behaviour under reaction conditions by introducing an inactive (but not denatured) intermediate in rapid equilibrium with the active form. The temperature midpoint (T-eq) of the rapid equilibration between the two forms is related to the growth temperature of the organism, and the enthalpy of the equilibrium (Delta H-eq) to its ability to function over various temperature ranges. In the present study, we show that the difference between the active and inactive forms is at the enzyme active site. The results reveal an apparently universal mechanism, independent of enzyme reaction or structure, based at or near, the active site, by which enzymes lose activity as temperature rises, as opposed to denaturation which is global. Results show that activity losses below T-eq may lead to significant errors in the determination of Delta G(cat)* made on the basis of the two-state ('Classical') model, and the measured k(cat) will then not be a true indication of an enzyme's catalytic power. Overall, the results provide a molecular rationale for observations that the active site tends to be more flexible than the enzyme as a whole, and that activity losses precede denaturation, and provide a general explanation in molecular terms for the effect of temperature on enzyme activity.