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

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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.