Abstract
Chloromethane (CM) is an ozone-depleting gas, produced predominantly from natural sources, that provides an important carbon source for microbes capable of consuming it. CM catabolism has been difficult to study owing to the challenging genetics of its native microbial hosts. Since the pathways for CM catabolism show evidence of horizontal gene transfer, we reproduced this transfer process in the laboratory to generate new CM-catabolizing strains in tractable hosts. We demonstrate that six putative accessory genes improve CM catabolism, though heterologous expression of only one of the six is strictly necessary for growth on CM. In contrast to growth of Methylobacterium strains with the closely related compound dichloromethane (DCM), we find that chloride export does not limit growth on CM and, in general that the ability of a strain to grow on DCM is uncorrelated with its ability to grow on CM. This heterologous expression system allows us to investigate the components required for effective CM catabolism and the factors that limit effective catabolism after horizontal transfer.
Highlights
Chloromethane (CM) is the most abundant organohalide on earth, accounting for roughly 16% of tropospheric chlorine in 2012, and contributes to chloride-catalyzed ozone depletion (World Meteorological Organization, 2014)
An abundant electron-rich compound represents a valuable carbon source for a microbe and, as expected, multiple microbial strains have been isolated based on their ability to grow with CM as the sole source of carbon and Horizontal Transfer of Chloromethane Catabolism energy (Hartmans et al, 1986; Doronina et al, 1996; McAnulla et al, 2001; Woodall et al, 2001; Schäfer et al, 2005; Warner et al, 2005; Nadalig et al, 2011; Nadalig et al, 2014)
Methylene-H4F is formed during conventional methylotrophic growth, wherein M. extorquens oxidizes methanol to formate in a tetrahydromethanopterin (H4MPT)-dependent pathway (Chistoserdova et al, 1998; Marx et al, 2003)
Summary
Chloromethane (CM) is the most abundant organohalide on earth, accounting for roughly 16% of tropospheric chlorine in 2012, and contributes to chloride-catalyzed ozone depletion (World Meteorological Organization, 2014). We have previously analyzed the factors that limit the effectiveness of a horizontally transferred pathway for catabolism of dichloromethane (DCM), an industrial solvent that differs from CM by only a single chlorine (Michener et al, 2014a). It is unclear how general those factors would be, even for a closely related compound such as CM, since the pathways for catabolism of CM and DCM have different enzymology and metabolic consequences (Figure 1A). We have deliberately transferred the CM catabolic pathway into naïve Methylobacterium strains, generating new CM-utilizing microbes. We show that growth on CM is not limited by chloride export, in contrast to growth on DCM
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