Southeastern Section - 67th Annual Meeting - 2018

Paper No. 23-9
Presentation Time: 11:00 AM


JIANG, Nannan, Bredesen Center, University of Tennessee at Knoxville, 443 Greve Hall, 821 Volunteer Blvd, Knoxville, TN 37996, YAN, Jun, Key Laboratory of Pollution Ecology and Environmental Engineering, Institute of Applied Ecology, Chinese Academy of Sciences, Shenyang, 110016, China and LĂ–FFLER, Frank E., Department of Microbiology, University of Tennessee at Knoxville, Knoxville, TN 37996

The biogeochemistry of chlorine is closely linked to the occurrence of chlorinated organic compounds, which require key cofactors for degradation. Corrinoids such as vitamin B12 are essential cofactors for all domains of life on our planet. These molecules have been shown to participate in DNA synthesis, methylation events, and dechlorination reactions. In the specific case of dechlorination, vitamin B12 can serve as cofactors for reductive dehalogenases (RDases) that catalyze hydrogenolytic carbon-halogen bond cleavage. The lower bases attached to corrinoid cofactors control the activity of RDases differently in corrinoid auxotrophic Dehalococcoides mccartyi (Dhc) strains, which prefer a corrinoid with 5,6-dimethylbenzimidazole (DMB) as the lower base. Geobacter sulfurreducens produces a corrinoid with 5-hydroxybenzimidazole (5-OHBza) as the lower base. 5-OHBza is a DMB precursor but does not support RDase activity in Dhc strains. Here, we cloned and heterologously expressed a unique Geobacter lovleyi O-methyltransferase gene implicated in the methylation of 5-OHBza to 5-methoxybenzimidazole (5-OMeBza). Distinct from 5-OHBza, 5-OMeBza supports Dhc reductive dechlorination activity. HPLC and LC-MS analyses demonstrated that the G. sulfurreducens mutant expressing the O-methyltransferase converted the native 5-OHBza-cobamide to the 5-OMeBza-cobamide. Co-cultivation of Dhc and the recombinant G. sulfurreducens in the absence of exogenous corrinoid supported Dhc growth and the complete dechlorination of cis-1,2-dichloroethene to ethene. Collectively, these studies demonstrated that this O-methyltransferase catalyzes a critical step in DMB biosynthesis and provides a genetic basis for the observation that Dhc reductive dechlorination activity occurs in co-cultures with G. lovleyi but not with G. sulfurreducens. These findings emphasize the critical role of the community for sustaining the activity of corrinoid-auxotrophic organohalide-respiring Chloroflexi, including Dhc. Since the anaerobic DMB biosynthesis pathway is limited to only a few organisms, cofactor metabolic engineering could provide feasible solutions for enhancing in-situ dechlorination activity and more efficiently restore sites contaminated by chlorinated compounds.