Density-functional investigation on the mechanism of H-atom abstraction by lipoxygenase
Density-functional investigation on the mechanism of H-atom abstraction by lipoxygenase
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DOI：
10.1007/s00775-002-0415-6
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发表时间：
2003-02-01
影响因子：
3
通讯作者：
Solomon, EI
中科院分区：
文献类型：
--
作者：
Lehnert, N;Solomon, EI
使用实验校准的密度泛函计算模型的大豆脂肪氧合酶1（SLO-1）的活性中心，洞察已获得铁活性中心的协调灵活性和其分子催化机制。SLO-1的亚铁形式显示出与弱配位Asn 694配体相关的溶液中配位数的变化。从计算中可以确定，弱Fe-O-694键与这种配位灵活性是由于Asn 694的侧向倾斜的几何形状，这是由蛋白质施加在该网站上。这种约束的释放（通过改变氢键网络）导致纯六配位位点。相比之下，酶的铁形式保持五配位。在这种情况下，配位水的去质子化在Asn 694的顺式位置提供强的羟基供体，从而削弱Fe-O-694键。因此，相对于氧化位点，Asn 694是还原位点的更强配体。利用这些实验标定的模型，计算了SLO-1中氢原子转移的反应能约为-18kcal/mol。所观察到的配位数从三价铁中的五配位到二价铁SLO-1中的六配位的变化增加了铁活性位点的还原电位。因此，蛋白质调节活性位点以获得最佳反应性。沿着反应坐标的电子结构分析表明，SLO-1中的H原子转移实际上对应于质子耦合电子转移（PCET）。转移的电子不局限于质子上，而是在协同质子隧穿-电子隧穿（PTET）过程中直接从基底隧穿到铁活性位点。过渡态中共价连接的Fe-O-H-C桥降低了能垒，并为这种隧穿提供了有效的超交换途径。从计算中估计PTET过程的热势垒约为+15千卡/摩尔，包括零点能量校正。这对应于接近1 s（-1）的热反应速率k（therm）。相比之下，在这些条件下，质子隧穿的速率可以高达2x 10（9）s（-1）。
Using experimentally calibrated density functional calculations on models of the active site of soybean lipoxygenase 1 (SLO-1), insight has been obtained into the coordination flexibility of the iron active site and its molecular mechanism of catalysis. The ferrous form of SLO-1 shows a variation in coordination number in solution that is related to a weakly coordinating Asn694 ligand. From the calculations it is determined that the weak Fe-O-694 bond associated with this coordination flexibility is due to a sideways tilted geometry of Asn694 that is imposed on the site by the protein. Release of this constraint (by altering the hydrogen bonding network) leads to a pure six-coordinate site. In contrast, the ferric form of the enzyme stays five-coordinate. In this case, deprotonation of a coordinated water gives a strong hydroxo donor in the cis position to Asn694, weakening the Fe-O-694 bond. Hence, Asn694 is a stronger ligand to the reduced relative to the oxidized site. Using these experimentally calibrated models, the reaction energy for H-atom transfer in SLO-1 has been calculated to be about -18 kcal/mol. The observed change in coordination number going from five-coordinate in ferric to six-coordinate in ferrous SLO-1 increases the reduction potential of the iron active site. Hence, the protein adjusts the active site for optimal reactivity. Analysis of the electronic structure along the reaction coordinate shows that the H-atom transfer in SLO-1 actually corresponds to a proton-coupled electron transfer (PCET). The transferred electron does not localize on the proton, but tunnels directly from the substrate to the ferric active site in a concerted proton tunneling-electron tunneling (PTET) process. The covalently linked Fe-O-H-C bridge in the transition state lowers the energy barrier and provides an efficient superexchange pathway for this tunneling. The thermal barrier for the PTET process is estimated from the calculations to be about +15 kcal/mol including zero-point energy corrections. This corresponds to a thermal reaction rate of k(therm) approximate to 1 s(-1). In comparison, the rate of proton tunneling can be as high as 2x10(9) s(-1) under these conditions.