Chen Gong, Kong Xian, Zhu Jingying, Lu Diannan, Liu Zheng
Department of Chemical Engineering, Tsinghua University, Beijing, 100084, China.
Phys Chem Chem Phys. 2015 Apr 28;17(16):10708-14. doi: 10.1039/c5cp00418g.
While the conjugation of enzymes with ABA copolymers has resulted in increased enzymatic activities in organic solvents, by several orders of magnitude, the underpinning mechanism has not been fully uncovered, particularly at the molecular level. In the present work, a coarse-grained molecular dynamics simulation of cytochrome c (Cyt c) conjugated with a PEO-PPO-PEO block copolymer (ABA) in toluene was simulated with Cyt c as a control. It is shown that the hydrophilic segments (PEO) of the conjugated block copolymer molecules tend to entangle around the hydrophilic patch of Cyt c, while the hydrophobic segments (PPO) extend into the toluene. At a lower temperature, the PEO tails tend to form a hairpin structure outside the conjugated protein, whereas the Cyt c-ABA conjugates tend to form larger aggregates. At a higher temperature, however, the PEO tails tend to adsorb onto the hydrophilic protein surface, thus improving the suspension of the Cyt c-ABA conjugates and, consequently, the contact with the substrate. Moreover, the temperature increase drives the conformational transition of the active site of Cyt c-ABA from an "inactive state" to an "activated state" and thus results in an enhanced activity. To validate the above simulations, Cyt c was conjugated to F127, an extensively used ABA copolymer. By elevating the temperature, a decrease in the average size of the Cyt c-F127 conjugates along with a great increase in the apparent activity in toluene was observed, as can be predicted from the molecular dynamics simulation. The above mentioned molecular simulations offer a molecular insight into the temperature-responsive behaviour of protein-ABA copolymers, which is helpful for the design and application of enzyme-polymer conjugates for industrial biocatalysis.
虽然酶与ABA共聚物的共轭作用已使酶在有机溶剂中的活性提高了几个数量级，但其潜在机制尚未完全揭示，尤其是在分子水平上。在本工作中，以细胞色素c（Cyt c）为对照，对其与聚环氧乙烷-聚环氧丙烷-聚环氧乙烷（PEO-PPO-PEO）嵌段共聚物（ABA）在甲苯中的共轭作用进行了粗粒度分子动力学模拟。结果表明，共轭嵌段共聚物分子的亲水性链段（PEO）倾向于缠绕在Cyt c的亲水性区域周围，而疏水性链段（PPO）则延伸到甲苯中。在较低温度下，PEO尾部倾向于在共轭蛋白外部形成发夹结构，而Cyt c-ABA共轭物则倾向于形成更大的聚集体。然而，在较高温度下，PEO尾部倾向于吸附在亲水性蛋白表面，从而改善Cyt c-ABA共轭物的悬浮性，并因此改善与底物的接触。此外，温度升高促使Cyt c-ABA活性位点的构象从“非活性状态”转变为“活化状态”，从而导致活性增强。为了验证上述模拟结果，将Cyt c与广泛使用的ABA共聚物F127进行共轭。如分子动力学模拟所预测的那样，通过升高温度，观察到Cyt c-F127共轭物的平均尺寸减小，同时在甲苯中的表观活性大幅增加。上述分子模拟为蛋白质-ABA共聚物的温度响应行为提供了分子层面的见解，这有助于设计和应用用于工业生物催化的酶-聚合物共轭物。
