Updating the Paradigm: Redox Partner Binding and Conformational Dynamics in Cytochromes P450.
Updating the Paradigm: Redox Partner Binding and Conformational Dynamics in Cytochromes P450.
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DOI：
10.1021/acs.accounts.1c00632
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发表时间：
2022-02-01
影响因子：
18.3
通讯作者：
Follmer AH
中科院分区：
文献类型：
--
作者：
Poulos TL;Follmer AH
本文综述了氧化还原伙伴结合、变构和构象动力学在细胞色素P450质子耦合电子转移中所起作用的最新研究结果。P450是自然界最大的酶家族之一，在需要底物氧化形成对生物体生命至关重要的关键分子或外源解毒时，P450并不少见。P450可以在非常大范围的底物上工作，从很小到很大，但整个P450的三维结构是保守的。考虑到结构的这种守恒性，一般认为基本催化机理是保守的。在几乎所有的P450中，O2的O-O键必须被异源裂解，使一个氧原子，即远端的氧，以水的形式离开，留下一个血红素铁连接的O原子作为强大的氧化剂，用于激活附近的底物。为了有效地进行这一过程，需要外部提供的电子和质子。当电子从通常包含Fe2S2或FMN氧化还原中心的氧化还原伙伴转移时，必须向离开的O原子添加两个质子。用来解开这些机制细节的范例P450一直是细菌的CYP101A1或P450cam。P450cam是针对其自身的Fe2S2氧化还原伙伴putidaredosin或PDX的，长期以来一直认为PDX可能通过将P450cam转换为活性构象而发挥效应/变构作用。P450cam-PDX络合物的晶体结构、光谱数据和直接结合实验提供了一些答案。PDX将P450cam的构象转变为更开放的状态，这一转变被认为是触发O2激活所需的质子中继网络。这个质子中继网络的一个重要部分是一个高度保守的天冬氨酸(有时是谷氨酸)，它是已知在许多P450中活动的关键。这种天冬氨酸和质子递送网络如何连接到氧化还原伙伴结合非常简单。在关闭状态下，这种天冬氨酸被盐桥束缚，但当PDX结合时，这些盐桥被破坏，使天冬氨酸自由发挥其在质子转移中的作用。另一种假设表明，一个特定的质子中继网并不是真正必要的。在这种情况下，天冬氨酸在打开/关闭转变中起结构作用，仅仅打开活性中心访问通道就足以使溶剂质子进入O2质子化。旨在测试这些不同假设的实验显示，P450 cam和其他细菌的P450都存在一些令人惊讶的现象。分子动力学和结晶学表明，P450cam可以经历相当重要的构象体操，导致活性部位的大规模重组，需要多种顺式/反式脯氨酸异构化。研究还发现，X射线驱动底物羟化是更好地理解天冬氨酸及其周围残基在催化中所起作用的有用工具。在这里，我们总结了这些最新的结果，这些结果提供了一幅更动态的P450催化图景。
This Account summarizes recent findings centered on the role that redox partner binding, allostery, and conformational dynamics plays in cytochrome P450 proton coupled electron transfer. P450s are one of Nature’s largest enzyme families and it is not uncommon to find a P450 wherever substrate oxidation is required in the formation of essential molecules critical to the life of the organism or in xenobiotic detoxification. P450s can operate on a remarkably large range of substrates from the very small to the very large, yet the overall P450 three-dimensional structure is conserved. Given this conservation of structure, it is generally assumed that the basic catalytic mechanism is conserved. In nearly all P450s, the O2 O–O bond must be cleaved heterolytically enabling one oxygen atom, the distal oxygen, to depart as water and leave behind a heme iron-linked O atom as the powerful oxidant that is used to activate the nearby substrate. For this process to proceed efficiently, externally supplied electrons and protons are required. Two protons must be added to the departing O atom while an electron is transferred from a redox partner that typically contains either a Fe2S2 or FMN redox center. The paradigm P450 used to unravel the details of these mechanisms has been the bacterial CYP101A1 or P450cam. P450cam is specific for its own Fe2S2 redox partner, putidaredoxin or Pdx, and it has long been postulated that Pdx plays an effector/allosteric role by possibly switching P450cam to an active conformation. Crystal structures, spectroscopic data, and direct binding experiments of the P450cam–Pdx complex provide some answers. Pdx shifts the conformation of P450cam to a more open state, a transition that is postulated to trigger the proton relay network required for O2 activation. An essential part of this proton relay network is a highly conserved Asp (sometimes Glu) that is known to be critical for activity in a number of P450s. How this Asp and proton delivery networks are connected to redox partner binding is quite simple. In the closed state, this Asp is tied down by salt bridges, but these salt bridges are ruptured when Pdx binds, leaving the Asp free to serve its role in proton transfer. An alternative hypothesis suggests that a specific proton relay network is not really necessary. In this scenario, the Asp plays a structural role in the open/close transition and merely opening the active site access channel is sufficient to enable solvent protons in for O2 protonation. Experiments designed to test these various hypotheses have revealed some surprises in both P450cam and other bacterial P450s. Molecular dynamics and crystallography show that P450cam can undergo rather significant conformational gymnastics that result in a large restructuring of the active site requiring multiple cis/trans proline isomerizations. It also has been found that X-ray driven substrate hydroxylation is a useful tool for better understanding the role that the essential Asp and surrounding residues play in catalysis. Here we summarize these recent results which provide a much more dynamic picture of P450 catalysis.
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影响因子：
2.9
作者：
Batabyal D;Lewis-Ballester A;Yeh SR;Poulos TL
通讯作者：
Poulos TL
影响因子：
5.6
作者：
HASEMANN, CA;RAVICHANDRAN, KG;DEISENHOFER, J
通讯作者：
DEISENHOFER, J
影响因子：
4.8
作者：
Nagano, S;Tosha, T;Poulos, TL
通讯作者：
Poulos, TL
影响因子：
4.8
作者：
Glascock, MC;Ballou, DP;Dawson, JH
通讯作者：
Dawson, JH
DOI：
10.1111/j.1432-1033.1977.tb11847.x
发表时间：
1977-01-01
期刊：
EUROPEAN JOURNAL OF BIOCHEMISTRY
影响因子：
--
作者：
LANGE, R;BONFILS, C;DEBEY, P
通讯作者：
DEBEY, P