Aminopyrrolnitrin

Active site modeling of dimerization interface in combination with site-directed mutagenesis indicates that the electron in the PrnD Rieske oxygenase can be transferred by either of two pathways, one involving Asp183' and the other involving Asn180'. In addition, the overexpression of the isc operon involved in the assembly of iron-sulfur clusters increased the catalytic activity of PrnD in Escherichia coli by a factor of at least 4.

Pyrrolnitrin is a bacterial metabolite, served as a natural lead of agricultural fungicides. In a previous study, fenpiclonil was proven to inhibit the oxidative transformation of aminopyrrolnitrin to pyrrolnitrin, catalyzed by aminopyrrolnitrin oxidase (PrnD). This monooxygenase has an interesting catalytic activity of selective oxidation of aromatic amines, rather than aliphatic amines. However, its structural details are not well understood. In this study, various analogues of pyrrolnitrin were prepared to elucidate the structures of active site of PrnD through structure-activity relationships. In vivo pyrrolnitrin biosynthesis inhibition was determined with Burkholderia sp. O33 and Pseudomonas fluorescens Pf-5. Quantitative analysis of pyrrolnitrin and precursors indicates that 2,3-disubstituted phenyl at 3rd carbon and small substituents at 4th carbon of pyrrole are strictly required to give strong inhibitory effects. In addition, dissociable proton of pyrrole is also critical for inhibitory activity. Molecular simulation with homology-based PrnD model suggests a highly restricted conformational space in active site. The results may help more detailed understanding of this unusual enzyme. In addition, the information will be useful for the development of novel fungicide, compatible with pyrrolnitrin-producing bacterium.

Context: Conjugated linoleic acid (CLA) may optimize body composition, yet mechanisms underlining its benefits are not clear in humans. Objective: We aimed to reveal the CLA-induced changes in the plasma metabolome associated with body composition improvement and the predictive performance of baseline metabolome on intervention responsiveness. Methods: Plasma metabolome from overnight fasted samples at pre- and post-intervention of 65 participants in a 12-week randomized, placebo-controlled trial (3.2 g/day CLA vs 3.2 g/day sunflower oil) were analyzed using untargeted LC-MS metabolomics. Mixed linear model and machine learning were applied to assess differential metabolites between treatments, and to identify optimal panel (based on baseline conventional variables vs metabolites) predicting responders of CLA-derived body composition improvement (increased muscle variables or decreased adiposity variables) based on dual-energy x-ray absorptiometry. Results: Compared with placebo, CLA altered 57 metabolites (P < 0.10) enriched in lipids/lipid-like molecules including glycerophospholipids (n = 7), fatty acyls (n = 6), and sphingolipids (n = 3). CLA-upregulated cholic acid (or downregulated aminopyrrolnitrin) was inversely correlated with changes in muscle and adiposity variables. Inter-individual variability in response to CLA-derived body composition change. The areas under the curves of optimal metabolite panels were higher than those of optimal conventional panels in predicting favorable response of waist circumference (0.93 [0.82-1.00] vs 0.64 [0.43-0.85]), visceral adiposity index (0.95 [0.88-1.00] vs 0.58 [0.35-0.80]), total fat mass (0.94 [0.86-1.00] vs 0.69 [0.51-0.88]) and appendicular fat mass (0.97 [0.92-1.00] vs 0.73 [0.55-0.91]) upon CLA supplementation (all FDR P < 0.05). Conclusion: Post-intervention metabolite alterations were identified, involving in lipid/energy metabolism, associated with body composition changes. Baseline metabolite profiling enhanced the prediction accuracy for responsiveness of CLA-induced body composition benefits. Trial registration: ClinicalTrials.gov NCT03915808. Keywords: body composition; conjugated linoleic acid; metabolomics.