The study is aimed at identifying the modifications needed to re-engineer methyl-coenzyme M reductase (MCR), a key enzyme in the biological formation and anaerobic oxidation of methane, for shale gas activation and subsequent conversion to liquid fuel and high-value chemicals. The technology would facilitate on-site processing of shale gas methane as an alternative to either gas flaring or costly storage or transporting options.
Methyl-coenzyme M reductase (MCR) is the key enzyme in the biological formation and anaerobic oxidation of methane, a potent greenhouse gas and biofuel. MCR catalyzes the conversion of coenzyme B (CoB-SH) and methyl-coenzyme M (CH3-S-CoM) to the mixed heterodisulfide, CoB-S-S-CoM, and methane. Recently, anaerobic methanotrophic archaea (ANME) have been shown to catalyze the anaerobic oxidation of methane (AOM). AOM is thought to operate, at least in part, as the reverse of methanogenesis, with a homolog of MCR catalyzing the first step in the pathway, the activation of methane with CoB-S-S-CoM. Thus, there is great potential for the use of ANME in natural gas-to-liquid fuel conversion strategies. Unfortunately, no pure culture of an ANME has been obtained to date, due in large part to their exceptionally slow growth rates and their syntrophic association with sulfate-reducing bacteria, which limits their use in large-scale conversion processes. An attractive alternative is to engineer the AOM pathway and its key enzyme, MCR, into a more suitable microorganism for use in the bioconversion of natural gas. However, MCR cannot currently be produced in an active (nor activatable) form in a heterologous host. The project will address this deficiency by identifying and characterizing the genes and corresponding enzymes required for the production of mature holo MCR. The activity of MCR is critically dependent on the unique nickel-containing tetrapyrrole, coenzyme F430. In addition to housing coenzyme F430, the active site of MCR contains several unprecedented post-translational modifications (PTMs). The exact roles these PTMs play in MCR catalysis are unknown, as are the identities of the genes responsible for their formation. A comparative genomics investigation was utilized to identify several genes conserved in all methanogens that are excellent candidates to be involved in MCR maturation. The project will thus delineate the roles these genes play in the production of functionally active MCR by developing an expression system for the: 1) In vivo synthesis of coenzyme F430; 2) Investigation of MCR PTM; and 3) Heterologous production of holo MCR. In addition to facilitating the use of MCR in the production of renewable biomethane and the conversion of shale gas to liquid fuel and other high-value chemicals, the project will offer interdisciplinary training opportunities for undergraduate and graduate students, as well as educational enrichment opportunities for K-12 students.
The award is co-funded by the ENG Office of Emerging Frontiers and Multidisciplinary Activities.
