Domain motion coupled to radical catalysis in ornithine aminomutase

原子和分子含有被称为电子的带负电荷的小粒子。通常，原子或分子上的两个电子相互配对，从而降低它们的总能量并增加它们的稳定性。分子或原子上未配对的或孤的电子称为自由基。与自由基相关的能量增加导致它们不稳定，并与其他分子（例如氧或水）发生反应。在生物体中，这些异常自由基反应的目标可以破坏细胞成分，如DNA或蛋白质，导致癌症、衰老甚至死亡。人类，像所有生物一样，有专门的分子来执行特定的生物功能。其中一些功能是由专门设计的酶来执行的。有趣的是，研究发现越来越多的酶利用自由基进行不寻常的和非常困难的化学反应。其中一类酶利用维生素B12作为自由基的储存库。当底物与b12酶结合时，b12分子分裂成两个自由基。这允许一个激进的继电器系统随之而来；在催化反应过程中，孤电子会在酶腔中从一个分子跳到另一个分子。与所有使用自由基的酶一样，b12酶面临的两个重要挑战是(i)控制自由基产生的时间和（ii）控制自由基在酶腔内的反应性和轨迹。在催化循环过程中，酶必须在一系列高度同步的事件中引导活性自由基走向预定目标，同时尽量减少或防止自由基因与错误的分子或原子（即水、氧或酶本身）碰撞而自我熄灭。为了更好地理解一般的酶是如何控制自由基的游离的，我们将把重点放在一种特殊的酶上，称为鸟氨酸氨基交换酶。这种酶的独特之处在于它不仅含有维生素B12，还含有维生素B6。B6和B12都是监测催化循环中自由基传播的敏感探针，使鸟氨酸氨基交换酶成为详细研究酶介导的自由基化学的理想系统。我们最近确定了酶的分子结构（即酶的每个原子在三维空间中的位置）。从这些信息中，我们可以检查酶的独特结构特征，使它能够将自由基引导到预定的目标。作为这个项目的一部分，我们还将研究自由基对的距离和相对取向，通过一种技术，我们把酶放在一个大的磁场中。在这种特殊的物理状态下，自由基将充当微型磁铁，我们将能够获得它们的直接环境（即邻近原子）以及到第二个自由基的距离和方向的信息。我们还将在催化循环的不同阶段捕获酶，并使用上述技术确定酶如何以及在多大程度上改变其构象以实现生产性自由基繁殖。通过这项研究，我们将更好地了解酶如何控制和利用与能量相关的自由基，从而使化学上困难和能量上具有挑战性的反应得以进行。

Atoms and molecules contain small negatively charged particles called electrons. Typically, two electrons on an atom or molecule pair with one another, thereby lowering their overall energy and increasing their stability. An unpaired or lone electron on a molecule or atom is referred to as a radical. The increased energy associated with radicals cause them to be unstable and reactive towards other molecules, for example oxygen or water. Within a living organism, the targets for these aberrant radical reactions can damage cellular components like DNA or protein, leading to cancers, aging or even death. Humans, like all living things, have specialised molecules for carrying out specific biological functions. Some of these functions are executed by specifically designed enzymes. Interestingly, research is uncovering an ever increasing list of enzymes which use radicals to carry out unusual and very difficult chemical reactions. A certain class of these enzymes uses vitamin B12, which serves as a reservoir for radicals. The B12-molecule splits into two radicals when the substrate binds to the B12-enzyme. This permits a radical relay system to ensue; the lone electron will hop from one molecule to the next in the enzyme cavity during the course of the catalytic reaction. Two important challenges faced by B12-enzymes, as with all enzymes which use radicals, are (i) controlling the timing of radical production and (ii) controlling the reactivity and trajectory of radicals within the enzyme cavity. During the course of the catalytic cycle, the enzyme must direct the reactive radicals toward their intended targets in a series of highly synchronized events, whilst at the same time minimize or prevent the radical from extinguishing itself by collision with the wrong molecule or atom (i.e. water, oxygen, or the enzyme itself). To better understand how enzymes in general control the peregrination of radicals, we will focus on a particular enzyme, termed ornithine aminomutase. This enzyme is unique in that it not only contains vitamin B12, but also vitamin B6. Both B6 and B12 serve as sensitive probes for monitoring radical propagation during the catalytic cycle, making ornithine aminomutase an ideal system for detailed investigation of enzyme-mediate radical chemistry. We have recently determined the molecular architecture of the enzyme, (i.e. where each atom of the enzyme is located in a three dimensional space). From this information, we can examine the unique structural features of the enzyme that enables it to direct radicals towards their intended target. As part of this programme we will also investigate the distance and relative orientation of the radical pair, by a technique whereby we place the enzyme in a large magnetic field. Under this particular physical state, the radicals will act as miniature magnets, and we will be able to derive information on their immediate environment, (i.e. neighbouring atoms) as well as the distance and orientation to a second radical. We will also trap the enzyme at different stages of the catalytic cycle, and use the above techniques to determine how and to what extent the enzyme changes its conformation to enable productive radical propagation. From this research, we will better understand how enzymes control and harness the energy associated radicals, enabling chemically difficult and energetically challenging reactions to be performed.