Mechanisms of Energy Conservation in Bifurcating Electron Transfer Flavoproteins

就像电线把电力输送到我们家里的每个房间一样，细胞里也有专门的蛋白质，把电子从产生电子的反应中输送到需要电子的重要反应中。该项目研究了一种新发现的“电子转移黄蛋白”（Etfs），它作为能量经纪人，通过接受低能量电子对并将其能量集中在一对电子上产生一个高能电子，以数量换取质量。至关重要的是，这个生化“升级站”使细胞能够从空气中固定氮气，从而产生自己的肥料，从而使原本无法生产粮食的地方成为可能。该项目旨在了解这些Etfs是如何完成这一复杂过程的。该研究试图通过研究发生这种情况的分子（这种黄素是核黄素维生素B2的衍生物）来了解能量在两个电子之间重新分配的位置的特殊之处。该研究还试图确定构建在蛋白质支架中的安全机制，使这种苛刻的转化在良性、无毒的蛋白质中发生。因此，这项工作旨在阐明该过程的基本原理，以便将其设计成人造设备和材料，以提高我们使用太阳能的能力，并提高我们使用电能的效率和多功能性。黄素只是众多植物色素中的一个例子，它们具有强大的化学特性和绚丽的色彩。项目科学家通过一系列研讨会和植物色素、纤维和香料课程分享他们对色素的热情，参与者在学习植物颜色的化学原理的同时，使用天然材料创作纤维艺术项目。这门创造性的化学课程让所有参与者都有机会设计实验，并将科学与艺术结合起来。一个长期熟悉的蛋白质家族最近被发现具有惊人的能力，可以在从NADH获得的电子对之间重新分配能量，产生比NADH源更具还原能力的电子（电子转移分岔，或“分岔”）。本研究确定了在新纯化的分叉电子转移黄蛋白（Etf）中确定的两种黄素中的哪一种发挥了不同的作用，然后试图了解蛋白质环境的哪些方面负责每种不同的活动（传统电子转移与分叉）。光谱电化学滴定测量单个氨基酸改变含有氨基酸取代的蛋白质中黄素还原电位（Eo）的程度，光谱研究阐明了黄素共价键、氢键和质子化状态的变化，绘制了活性位点在产生分岔活性中的作用。目标残基包括与固氮相关的所有Etfs共有但在其他Etfs中不保守的半胱氨酸。因此，这项研究可能揭示了在抑制副作用的同时利用黄素的反应能力所需的蛋白质背景。目的是阐明原理的多样性和效率的分岔，实施超越生物化学。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

Just as electrical wires carry power to every room in our houses, cells have dedicated proteins carrying a current of electrons from reactions that generate electrons to vital reactions that require them. This project addresses a newly-recognized class of 'electron transferring flavoproteins' (Etfs) that act as energy brokers, trading quantity for quality by accepting pairs of modest-energy electrons and concentrating their energy onto just one of the pair to produce one high-energy electron. Crucially, this biochemical 'step-up station' makes it possible for cells to fix nitrogen gas from air to generate their own fertilizer, making food production possible where it otherwise would not be. The project seeks to understand HOW these Etfs accomplish this complex process. The research seeks to learn what is special about the site at which energy is reallocated among the two electrons by studying the molecule on which this happens (this flavin is a derivative of the riboflavin vitamin B2). The research also seeks to identify safety mechanisms built into the protein scaffold that allow such demanding transformations to occur within a benign, non-toxic protein. Thus the work aims to articulate the underlying principles of the process, so that they can be designed into human-made devices and materials, to increase our ability to use solar power and boost the efficiency and versatility with which we use electrical energy in general. Flavins are just one example of many plant pigments that present powerful chemical properties along with glorious colors. The project scientists share their enthusiasm for pigments via a series of workshops and a course on plant pigments, fibers and fragrances in which participants create a fiber art project using natural materials while learning about the chemical principles underlying plant colors. This creative chemistry course gives all participants a chance to design experiments and integrate science and art.A long-familiar family of proteins has recently been found to have a surprising capacity to re-allocate energy among pairs of electrons acquired from NADH, yielding one with more reducing power than the NADH source (electron transfer bifurcation, or 'bifurcation'). This research establishes which of the two flavins plays which of the contrasting roles identified in the newly-purified bifurcating electron transferring flavoprotein (Etf), and then seeks to understand what aspects of the protein environment are responsible for each of the different activities (conventional electron transfer vs. bifurcation). Spectroelectrochemical titrations measure the extent to which individual amino acids change the flavin reduction potentials (Eo) in proteins bearing amino acid substitutions, and spectroscopic studies elucidate changes in flavin covalent bonding, hydrogen bonding and protonation states, mapping out the active site's role in producing bifurcating activity. Residues to be targeted include a cysteine common to all the Etfs associated with nitrogen fixation but not conserved in others. Thus the research may reveal the protein context needed in order to exploit flavins' capacity for this reactivity while suppressing side-effects. The objective is to elucidate principles underlying the versatility and efficiency of bifurcation, for implementation beyond biochemistry.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.