Hot spots in cold adaptation:: Localized increases in conformational flexibility in lactate dehydrogenase A4 orthologs of Antarctic notothenioid fishes
Hot spots in cold adaptation:: Localized increases in conformational flexibility in lactate dehydrogenase A4 orthologs of Antarctic notothenioid fishes
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DOI：
10.1073/pnas.95.19.11476
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发表时间：
1998-09-15
影响因子：
11.1
通讯作者：
Somero, GN
中科院分区：
文献类型：
--
作者：
Fields, PA;Somero, GN
为了阐明酶对极端寒冷的适应机制，我们测定了9个南极(-1.86~1℃)和3个南美洲(4~10℃)拟脊椎动物的乳酸脱氢酶A(4)(A(4)-LDH)的动力学性质、热稳定性和推导的氨基酸序列，较高的Michaelis-Menten常数(K-m)和催化速率常数(k(CAT))区分了南极和南美物种的直系物，但适应温度与由于热变性而失去活性的速率之间没有关系。在所有物种中，活性中心残基都是完全保守的，k(CAT)和K-m的差异是由分子中其他位置的取代引起的。在地理群体内，不同的替代产生相同的动力推进力。通过将我们的数据与其他脊椎动物的A(4)-LDH序列相结合，以及关于局部构象变化在设定k(CAT)中所起的作用的信息，我们得出结论，Nototheniid A(4)-LDH通过增加分子小区域的灵活性来适应寒冷的温度，这影响了相邻活性部位结构的移动性，利用这些发现，我们提出了一个模型，解释了K-m和k(CAT)的温度适应性变化。序列的改变增加了催化构象变化所涉及的酶区域的灵活性，这可能会减少构象中这些速率控制变化的能量(焓)障碍，从而增加k(CAT)。然而，在普通的测量温度下，冷适应酶的较高构象熵可能会促进与配体结合较差的构象，从而导致相对于热适应同源酶的高K-m值。
To elucidate mechanisms of enzymatic adaptation to extreme cold, we determined kinetic properties, thermal stabilities, and deduced amino acid sequences of lactate dehydrogenase A(4) (A(4)-LDH) from nine Antarctic (-1.86 to 1 degrees C) and three South American (4 to 10 degrees C) notothenioid teleosts, Higher Michaelis-Menten constants (K-m) and catalytic rate constants (k(cat)) distinguish orthologs of Antarctic from those of South American species, but no relationship exists between adaptation temperature and the rate at which activity is lost because of heat denaturation. In all species, active site residues are conserved fully, and differences in k(cat) and K-m are caused by substitutions elsewhere in the molecule. Within geographic groups, identical kinetic propel-ties are generated by different substitutions. By combining our data with A(4)-LDH sequences for other vertebrates and information on roles played by localized conformational changes in setting k(cat), we conclude that notothenioid A(4)-LDHs have adapted to cold temperatures by increases in flexibility in small areas of the molecule that affect the mobility of adjacent active-site structures, Using these findings, we propose a model that explains linked temperature-adaptive variation in K-m and k(cat). Changes in sequence that increase flexibility of regions of the enzyme involved in catalytic conformational changes may reduce energy (enthalpy) barriers to these rate-governing shifts in conformation and, thereby, increase k(cat). However, at a common temperature of measurement, the higher configurational entropy of a cold-adapted enzyme may foster conformations that bind ligands poorly, leading to high K-m values relative to warm-adapted orthologs.