A metabolomic view of how the human gut microbiota impacts the host metabolome using humanized and gnotobiotic mice.

Defining the functional status of host-associated microbial ecosystems has proven challenging owing to the vast number of predicted genes within the microbiome and relatively poor understanding of community dynamics and community-host interaction. Metabolomic approaches, in which a large number of small molecule metabolites can be defined in a biological sample, offer a promising avenue to 'fingerprint' microbiota functional status. Here, we examined the effects of the human gut microbiota on the fecal and urinary metabolome of a humanized (HUM) mouse using an optimized ultra performance liquid chromatography-mass spectrometry-based method. Differences between HUM and conventional mouse urine and fecal metabolomic profiles support host-specific aspects of the microbiota's metabolomic contribution, consistent with distinct microbial compositions. Comparison of microbiota composition and metabolome of mice humanized with different human donors revealed that the vast majority of metabolomic features observed in donor samples are produced in the corresponding HUM mice, and individual-specific features suggest 'personalized' aspects of functionality can be reconstituted in mice. Feeding the mice a defined, custom diet resulted in modification of the metabolite signatures, illustrating that host diet provides an avenue for altering gut microbiota functionality, which in turn can be monitored via metabolomics. Using a defined model microbiota consisting of one or two species, we show that simplified communities can drive major changes in the host metabolomic profile. Our results demonstrate that metabolomics constitutes a powerful avenue for functional characterization of the intestinal microbiota and its interaction with the host.

Structural characterization of enterobactin hydrolase IroE.

The proliferation of many pathogenic bacteria is limited by the scarcity of soluble iron in their environment. Many of these bacteria scavenge iron by synthesizing and exporting small molecule siderophores that chelate iron. Iron-bound siderophores are subsequently imported for metabolic processing. Three related serine hydrolases have been characterized biochemically in this pathway: Fes, IroD, and IroE. Here, we report the crystal structure of IroE from uropathogenic Escherichia coli CFT073. The native structure and a complex with diisopropyl fluorophosphonate (DFP, a potent serine hydrolase inhibitor) were determined at 2.3 and 1.4 A resolution, respectively. IroE has the typical alpha/beta-hydrolase fold with an atypical catalytic dyad composed of Ser 189 and His 287. Mutation of either residue was detrimental to catalysis. In addition, rather than the typical oxyanion hole composed of backbone amides, IroE employs the atypical guanidinium moiety of Arg 130. Asp 90 anchors Arg 130 in the active site, and mutation of either residue was likewise detrimental to catalysis. We also compare the structure of IroE to the structure of Fes from Shigella flexneri (PDB entry 2B20). Both enzymes have similar active sites, but Fes has an additional amino-terminal lid domain. These lid domains are proposed to confer specificity to these related hydrolases.