Bacterial phylotypes inhabiting the mucosa of small intestine and colon samples from CM and RM mice identified by high throughput Illumina analysis of the 16S rRNA gene.

<p>Bacterial phylotypes inhabiting the mucosa of small intestine and colon samples from CM and RM mice identified by high throughput Illumina analysis of the 16S rRNA gene.</p

Urine metabolomics in RM and CM mouse models.

<p>Panel A and C. Volcano plots facilitating visualization of differentially abundant metabolites that were selected based on fold change (X-axis) and p-value in (Y-axis) for Atm-KO and WT mice, respectively. The m/z values highlighted in pink have a fold change of ≥ 0.5 or ≤ 2.0 and p-value ≤ 0.05 in RM as compared to CM mice and were selected for further characterization. Panel B and D: PCA plots showing separation between RM and CM in Atm-KO and WT mice, respectively.</p

Gut microbiota modulates fecal metabolomic profiles in Atm-KO and WT mice.

<p>Panel A and C. Volcano plots facilitating visualization of differentially abundant metabolites that were selected based on fold change (X-axis) and p-value in (Y-axis) for Atm-KO and WT mice, respectively. The m/z values highlighted in pink have a fold change of ≥ 0.5 or ≤ 2.0 and p-value ≤ 0.05 in RM as compared to CM mice and were selected for further characterization. Panels B and D: PCA plots showing separation between RM and CM in Atm-KO and WT mice, respectively.</p

Functional Pathway Analysis showing major upregulated pathways in Atm-KO mice with RM as compared to CM microbiota.

<p>Panel A shows significantly perturbed canonical pathways in Atm-KO-RM mice, while Panel B shows a TP53 regulated network that correlated strongly with metabolic profiles of Atm-KO-RM mice.</p

Heat map showing differential abundance of urine metabolites in the various study groups: Metabolites profiles for Atm-KO-CM group, Atm-KO-RM, WT-CM, and WT-RM, respectively.

<p>Each column represents a sample, and each row represents a metabolite. The mean signal intensity of CM group is colored black; red indicates above-mean intensity, green denotes below mean intensity, and the degree of color saturation reflects the magnitude of intensity relative to the mean.</p

Fecal metabolites that were differentially abundant in RM as compared to CM only in WT mice.

<p>Fecal metabolites that were differentially abundant in RM as compared to CM only in WT mice.</p

Heat map showing differential abundance of fecal metabolites in various study groups (Panels A-D): Metabolites profiles for Atm-KO-CM group, Atm-KO-RM, WT-CM, and WT-RM, respectively.

<p>Each column represents a sample, and each row represents a metabolite. The mean signal intensity of CM group is colored black; red indicates above-mean intensity, green denotes below mean intensity, and the degree of color saturation reflects the magnitude of intensity relative to the mean.</p

Urine metabolites that were differentially abundant in RM as compared to CM in only Atm-KO mice.

<p>Urine metabolites that were differentially abundant in RM as compared to CM in only Atm-KO mice.</p

Fecal metabolites that were differentially abundant in RM as compared to CM only in Atm-KO mice.

<p>Fecal metabolites that were differentially abundant in RM as compared to CM only in Atm-KO mice.</p

Kinetic Characterization and Allosteric Inhibition of the <i>Yersinia pestis</i> 1-Deoxy-D-Xylulose 5-Phosphate Reductoisomerase (MEP Synthase)

<div><p>The methylerythritol phosphate (MEP) pathway found in many bacteria governs the synthesis of isoprenoids, which are crucial lipid precursors for vital cell components such as ubiquinone. Because mammals synthesize isoprenoids via an alternate pathway, the bacterial MEP pathway is an attractive target for novel antibiotic development, necessitated by emerging antibiotic resistance as well as biodefense concerns. The first committed step in the MEP pathway is the reduction and isomerization of 1-deoxy-D-xylulose-5-phosphate (DXP) to methylerythritol phosphate (MEP), catalyzed by MEP synthase. To facilitate drug development, we cloned, expressed, purified, and characterized MEP synthase from <i>Yersinia pestis</i>. Enzyme assays indicate apparent kinetic constants of K_M^DXP = 252 µM and K_M^NADPH = 13 µM, IC₅₀ values for fosmidomycin and FR900098 of 710 nM and 231 nM respectively, and K_i values for fosmidomycin and FR900098 of 251 nM and 101 nM respectively. To ascertain if the <i>Y. pestis</i> MEP synthase was amenable to a high-throughput screening campaign, the Z-factor was determined (0.9) then the purified enzyme was screened against a pilot scale library containing rationally designed fosmidomycin analogs and natural product extracts. Several hit molecules were obtained, most notably a natural product allosteric affector of MEP synthase and a rationally designed bisubstrate derivative of FR900098 (able to associate with both the NADPH and DXP binding sites in MEP synthase). It is particularly noteworthy that allosteric regulation of MEP synthase has not been described previously. Thus, our discovery implicates an alternative site (and new chemical space) for rational drug development.</p></div