Comparative genomics of ethanolamine utilization

Abstract

Ethanolamine can be used as a source of carbon and nitrogen by phylogenetically diverse bacteria. Ethanolamine-ammonia lyase, the enzyme that breaks ethanolamine into acetaldehyde and ammonia, is encoded by the gene tandem eutBC. Despite extensive studies of ethanolamine utilization in Salmonella enterica serovar Typhimurium, much remains to be learned about EutBC structure and catalytic mechanism, about the evolutionary origin of ethanolamine utilization, and about regulatory links between the metabolism of ethanolamine itself and the ethanolamine-ammonia lyase cofactor adenosylcobalamin. We used computational analysis of sequences, structures, genome contexts, and phylogenies of ethanolamine-ammonia lyases to address these questions and to evaluate recent data-mining studies that have suggested an association between bacterial food poisoning and the diol utilization pathways. We found that EutBC evolution included recruitment of a TIM barrel and a Rossmann fold domain and their fusion to N-terminal alpha-helical domains to give EutB and EutC, respectively. This fusion was followed by recruitment and occasional loss of auxiliary ethanolamine utilization genes in Firmicutes and by several horizontal transfers, most notably from the firmicute stem to the Enterobacteriaceae and from Alphaproteobacteria to Actinobacteria. We identified a conserved DNA motif that likely represents the EutR-binding site and is shared by the ethanolamine and cobalamin operons in several enterobacterial species, suggesting a mechanism for coupling the biosyntheses of apoenzyme and cofactor in these species. Finally, we found that the food poisoning phenotype is associated with the structural components of metabolosome more strongly than with ethanolamine utilization genes or with paralogous propanediol utilization genes per se.

FIG. 1.

Diversity of eutBC genome contexts. Short operons: A, Deltaproteobacteria; B, a subset of Proteobacteria, Chlorophlexi, and Bacteroidetes; C, selected Proteobacteria and Acidobacteria; and D, Betaproteobacteria (eutR is in a different genomic location than eutBC and eat). Long operons: E, Nocardioides sp.; F, Enterobacteriaceae; G, M. aquaeolei; H, S. boydii Sb227; I, S. sonnei Ss046; J, S. dysenteriae Sd197; K, Symbiobacterium thermophilum and P. luminescens; L, P. fluorescens Pf-5; M, Clostridiaceae and F. nucleatum; N, Listeriaceae and Enterococcaceae; and O, C. acetobutylicum. A probable eutB pseudogene is shown in white. The predicted EutR-binding sites are indicated by purple ellipses. See Tables S1 and S2 in the supplemental material for complete lists of gene names and species.

FIG. 2.

Maximum likelihood evolutionary tree of EutB. The bootstrap support of tree partitions is indicated by branch color: green, >70%; blue, 50 to 70%; and red, <50%. The outer color stripe mark genes from the short operon in red and genes from the long operon in green. The inner color circle marks bacterial clades: red, Alphaproteobacteria; orange, Betaproteobacteria; green, Gammaproteobacteria; dark blue, Deltaproteobacteria; dark purple, Firmicutes; pink, Actinobacteria; lime, Acidobacteria; light purple, Fusobacteria; light blue, Chlorophlexi; and light green, Bacteroidetes. The shaded background marks two clades that do not agree with established bacterial phylogeny and suggest horizontal gene transfer events (see text).

FIG. 3.

Conserved elements in proteobacteria that may bind EutR. (A) Conserved element preceding the eut operon in Betaproteobacteria (for site scores and locations, see Table S3 in the supplemental material); (B) conserved binding element preceding the eut operon in Enterobacteriaceae; (C) conserved element upstream of the cbiA gene in Enterobacteriaceae.