Statement of Project Objectives The technology described in this proposal is a radical departure from gas-to-liquid (GTL) technologies based on native methane monooxygenases (MMOs). Instead of beginning with a complex, multisubunit MMO, the primary objective of this proposal is to redesign a well-characterized multicopper oxidase (MCO) to perform methane-oxidation chemistry within the confines of single protein subunit. This scaffold will be tractable for protein engineering efforts, and will be employable in a wide variety of applications. Development of this technology will first focus on creating a robust and versatile protein scaffold (Task 1) that performs MMO chemistry (Task 2). Once MMO chemistry has been developed, we will engineer enzymes that catalyze highly efficient methane oxidation chemistry without the input of reductant (Task 3), and enzymes that catalyze methane C-C coupling (Task 4). Tasks 2 and 3 will result in versatile high-turnover and high-efficiency methanol-producing enzymes that can replace traditional MMOs in biological fuel-production pathways. Task 4 will result in enzymes capable of methane C-C coupling that can be further engineered to produce unique GTL products without biological assimilation of methanol. Tasks 1 and 2 will lay the foundation for further engineering and application development. In Task 1, we will design MCO variants that recapitulate the structural and redox features of the metal centers that enable catalysis in native MMOs. In Task 2, we will combine these individual cofactors into one protein, demonstrate methane-oxidation activity, and perform additional rounds of engineering to optimize this activity. Tasks 3 and 4 will build on Tasks 2 and 3, respectively, to develop biologically unprecedented chemistries that enable the highly efficient conversion of natural gas to liquid fuels. Given that Task 1 and Task 2 are critical paths, we have assigned ~35% effort each to Tasks 1 and 2 and ~15% effort each to Tasks 3 and 4.
|Effective start/end date||2/12/14 → 11/15/15|
- Advanced Research Projects Agency - Energy (DE-AR0000435/0004)
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