Collaborative Research: Synthetic methane fixation cascades based on engineered membrane vesicles for biofuel cell applications

石油和天然气开采技术的进步使得主要由甲烷组成的天然气广泛可用。在这些操作过程中，大量甲烷泄漏到大气中。井场往往偏远和孤立。在这些情况下，标准的捕获和处理技术一般不可行。 该项目的目标是将甲烷转化为电力。 人工酶级联将被创造出来，将甲烷完全氧化为二氧化碳，并在此过程中产生电子。这些电子将用于燃料电池应用。 该项目将包括一个教育和推广计划，扩大学生获得基于项目的学习。开发一个综合的教学、研究和课程开发平台将吸引研究生、本科生和高中生，特别是那些代表性不足的群体。由于缺乏有效的酶级联，在环境温度下将大部分废弃的甲烷转化为更有价值的产品方面存在很大差距。为了解决这个问题，固定甲烷到甲醇耦合到甲醇氧化的概念，用于电子释放使用有组织的四酶级联膜囊泡。甲烷氧化细菌中的膜结合酶在温和条件下将甲烷选择性转化为甲醇。这是该方法的基础。采用新的混合酶合成生物学人工酶级联，使用直接从宿主膜分离的酶结合囊泡，应该完全氧化甲醇为CO2和产生电子的燃料电池应用。具体地，将蛋白质支架缀合到甲烷固定酶结合的膜囊泡上将使得能够组装三种脱氢酶，用于将甲烷顺序转化为二氧化碳。这三种酶与甲烷固定酶的共定位将促进酶之间由于底物通道而产生的协同作用，从而导致总电流密度的增强。级联酶的一个附加益处是能够改善脱氢酶和甲烷固定酶之间的NADH和NAD+转移，以获得更高的催化效率。预期的结果是一个新的平台，产生合成膜囊泡系统，用于甲烷固定到CO2为生物燃料电池提供动力，并作为其他膜结合酶系统的技术平台。该奖项反映了NSF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。

Advances in oil and gas extraction techniques have made natural gas, composed primarily of methane, widely available for use. Large quantities of methane leak into the atmosphere during these operations. Well sites are often remote and isolated. Standard capture and treatment technologies are not generally feasible to apply in these cases. The objective of this project is to convert methane to electric power. Artificial enzyme cascades will be created to completely oxidize methane to CO2, generating electrons in the process. The electrons will be used in fuel cell applications. This project will include an education and outreach program that expands student access to project-based learning. Developing an integrated teaching, research, and curriculum development platform will engage graduate students, undergraduate, and high school students, particularly those in underrepresented groups.There is a big gap on converting mostly wasted methane to more valuable products under ambient temperature due to the lack of efficient enzyme cascades. To address this problem, the concept of fixing methane to methanol coupled to oxidation of methanol for electron release using an organized four-enzyme cascade on membrane vesicles is proposed. The membrane-bound enzyme found in methanotrophic bacteria will selectively convert methane to methanol under mild conditions. This is the foundation of the approach. Employing new hybrid enzyme-synthetic biology artificial enzyme cascades, using the enzyme-bound vesicles isolated directly from the host membranes, should completely oxidize methanol to CO2 and generate electrons for fuel-cell applications. Specifically, conjugating a protein scaffold onto the methane fixing enzyme-bound membrane vesicles will enable the assembly of three dehydrogenases for the sequential conversion of methane to carbon dioxide. Co-localization of the three dehydrogenases with the methane fixing enzyme will promote the synergistic action between the enzymes due to substrate channeling, resulting in an enhancement in the overall current density. One added benefit of cascading the enzymes is to enable the improved transfer of NADH and NAD+ between dehydrogenase and the methane fixing enzyme for higher catalytic efficiency. The expected result is a new platform to generate synthetic membrane vesicles system for methane fixation to CO2 to power biofuel cells, and to serve as a technology platform for other membrane-bound enzyme systems.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.