Mechanistic Insights into Substrate Positioning That Distinguish Non-heme Fe(II)/α-Ketoglutarate-Dependent Halogenases and Hydroxylases
Mechanistic Insights into Substrate Positioning That Distinguish Non-heme Fe(II)/α-Ketoglutarate-Dependent Halogenases and Hydroxylases
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DOI：
10.1021/acscatal.2c06241
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发表时间：
2023-02-03
期刊：
影响因子：
12.9
通讯作者：
Kulik, Heather J.
中科院分区：
文献类型：
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作者：
Kastner, David W.;Nandy, Aditya;Kulik, Heather J.
生理条件下不同生物产物的非血红素铁卤化酶和羟化酶作用。为了更好地理解底物定位的差异是其不同反应性的关键，我们汇编了可用的晶体学和光谱数据，这些数据显示羟化酶更喜欢锐角氧代-Fe-H靶角，而卤化酶更喜欢钝角。通过分子动力学模拟，我们模拟了具有代表性的羟化酶TauD和VioC和卤化酶BesD和WelO 5与急性和钝谐波约束。我们确定了关键的底物相互作用伙伴，保持在各自的酶，如Asp 94在TauD和His 127在BesD的方法的角度。此外，我们的模拟结果表明，蛋白质环境中的卤化酶防止采样的锐角中观察到的羟化酶，反之亦然。为了验证这些经典的观察结果，我们优化了结构与大规模量子力学（QM）模拟，并确认QM衍生的底物酶氢键强度较高的本地配置。我们计算了限速氢原子转移步骤的反应势垒，发现无论酶-底物复合物如何，从锐角看反应势垒都略低。卤化酶反应坐标的分析揭示了形成的Fe（III）-羟基，单齿琥珀酸盐，和第二个协调领域，可以抑制羟基反弹的成员之间的氢键网络。
Non-heme iron halogenases and hydroxylases tion of diverse biological products under physiological conditions. To better understand the differences in substrate positioning key to their divergent reactivities, we compiled available crystallographic and spectroscopic data, which revealed that hydroxylases prefer an acute oxo-Fe-H target angle while halogenases prefer a more obtuse angle. With molecular dynamics simulations guided by this experimental information, we simulated the representative hydroxylases TauD and VioC and the halogenases BesD and WelO5 with both acute and obtuse harmonic restraints. We identified key substrate interaction partners that maintain the angle of approach in the respective enzymes, such as Asp94 in TauD and His127 in BesD. Moreover, our simulations reveal that the protein environment in halogenases prevents the sampling of acute angles observed in hydroxylases and vice versa. To validate these classical observations, we optimized the structure with large-scale quantum mechanical (QM) simulations and confirmed that QM-derived substrate-enzyme hydrogen bond strengths were higher in the native configurations. We computed reaction barriers for the rate-limiting hydrogen atom transfer step and found them to be slightly lower from an acute angle regardless of the enzyme-substrate complex. Analysis of the halogenase reaction coordinate reveals the formation of hydrogen bonding networks between the Fe(III)-hydroxyl, monodentate succinate, and a member of the second coordination sphere that may inhibit the hydroxyl rebound.