Abstract
Methylamine plays an important role in the global carbon and nitrogen budget; microorganisms that grow on reduced single carbon compounds, methylotrophs, serve as a major biological sink for methylamine in aerobic environments. Two non-orthologous, functionally degenerate routes for methylamine oxidation have been studied in methylotrophic Proteobacteria: Methylamine dehydrogenase and the N-methylglutamate pathway. Recent work suggests the N-methylglutamate (NMG) pathway may be more common in nature than the well-studied methylamine dehydrogenase (MaDH, encoded by the mau gene cluster). However, the distribution of these pathways across methylotrophs has never been analyzed. Furthermore, even though horizontal gene transfer (HGT) is commonly invoked as a means to transfer these pathways between strains, the physiological barriers to doing so have not been investigated. We found that the NMG pathway is both more abundant and more universally distributed across methylotrophic Proteobacteria compared to MaDH, which displays a patchy distribution and has clearly been transmitted by HGT even amongst very closely related strains. This trend was especially prominent in well-characterized strains of the Methylobacterium extroquens species, which also display significant phenotypic variability during methylamine growth. Strains like Methylobacterium extorquens PA1 that only encode the NMG pathway grew on methylamine at least five-fold slower than strains like Methylobacterium extorquens AM1 that also possess the mau gene cluster. By mimicking a HGT event through the introduction of the M. extorquens AM1 mau gene cluster into the PA1 genome, the resulting strain instantaneously achieved a 4.5-fold increase in growth rate on methylamine and a 11-fold increase in fitness on methylamine, which even surpassed the fitness of M. extorquens AM1. In contrast, when three replicate populations of wild type M. extorquens PA1 were evolved on methylamine as the sole carbon and energy source for 150 generations neither fitness nor growth rate improved. These results suggest that the NMG pathway permits slow growth on methylamine and is widely distributed in methylotrophs; however, rapid growth on methylamine can be achieved quite readily through acquisition of the mau cluster by HGT.
Highlights
The simplest methylated amine, mono-methylamine (CH3NH2; MA) is a toxic, inflammable organic compound that plays an important role in the carbon and nitrogen biogeochemical cycles [1,2,3,4,5,6,7] and contributes significantly to the biogenesis of greenhouse gases like methane [8,9]
The distribution of the NMG pathway and methylamine dehydrogenase (MaDH) in methylotrophic Proteobacteria that contain either one or both routes for MA oxidation are shown in have the NMG pathway and only 5 encode the mau gene cluster
In order to determine whether the patchy distribution of MaDH across sequenced methylotrophic Proteobacteria indicates transfer by frequent horizontal gene transfer (HGT), we constructed maximum-likelihood (ML) amino acid phylogenies for key genes of each MA oxidation module
Summary
The simplest methylated amine, mono-methylamine (CH3NH2; MA) is a toxic, inflammable organic compound that plays an important role in the carbon and nitrogen biogeochemical cycles [1,2,3,4,5,6,7] and contributes significantly to the biogenesis of greenhouse gases like methane [8,9]. In contrast to previous findings, recent studies with Methyloversatilis universalis FAM5 [15], Methylobacterium extorquens PA1 (referred to as PA1 from here on) (16) and various other bacteria [29,30] have shown that these strains can grow on MA despite lacking the mau gene cluster In these methylotrophs, MA oxidation is mediated by the NMG pathway that, unlike MaDH, requires three enzymatic steps and generates two amino acid derivatives as metabolic intermediates [31,32,33,34]. Whereas all M. extorquens strains possess the NMG pathway [21,37,38], AM1 and CM4 contain MaDH [21] (Figure 1A,B) These genotypic differences are known to influence MA growth: AM1 can grow on MA with rates that are five-fold higher than those observed for PA1 [16,38]. We directly demonstrate that an HGT event can instantaneously overcome physiological constraints imposed by certain metabolic pathways and dramatically change the performance and fitness of an organism on a growth substrate
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