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Aims Propolis is a resinous bee product containing several hundred biologically active compounds. Although the antibacterial activity of propolis has been demonstrated in many in vitro studies, less is known about its mode of action. In this study, we aimed to shed some light on the antibacterial mechanism of action of propolis against Escherichia coli BW25113 using a nuclear magnetic resonance (NMR) based metabolomics approach. Methods E. coli BW25113 cells were subjected to different sub-lethal concentrations (0, 2, 4, and 6 mg/mL) of Turkish propolis water extract (PWE). The 500-MHz 1H NMR spectroscopy was then employed to ascertain the metabolic profiles of E. coli extracts. Results A total of 52 metabolites were identified from the NMR spectra, belonging to 17 main classes, such as amino acids and peptides, purines, and fatty acids. Twelve out of these 52 metabolites displayed remarkable changes at all tested PWE concentrations when compared to control conditions (P  < .05). Levels of 28 more metabolites were significantly altered in at least one of the three PWE treatments. The results of partial least squares discriminant analysis showed that there was a clear separation between control and propolis-treated cells and that putrescine, adenine, adenosine, guanosine, glucose, N6-acetyllysine, and acetamide had the highest effect on group differentiation. Finally, quantitative pathway analysis revealed that purine metabolism was significantly affected by PWE treatments. Conclusions Our results suggest that PWE inhibits the growth of E. coli BW25113 by affecting nucleic acid metabolism to a great extent. To the best of our knowledge, this is the first study to evaluate the global metabolic response of a bacterium to propolis. [ABSTRACT FROM AUTHOR]

Research has been conducted on the toluene 4-monooxygenase from Pseudomonas mendocina that catalyzes the NADH- and dioxygen-dependent toluene hydroxylation to form p-cresol. The solution structure of this monooxygenase has been investigated and the results are discussed.

Nonconservative mutations of the critical residues S47, F65, and E94, on the surface of ferredoxin have large parallel effects on the reduction potential and the electron-transfer reactivity of the (2Fe-2S) cluster. The reduction potential changes are not the principal factor governing electron transfer reactivity. The specific orientation of the proteins within the transient electron-transfer complex control the kinetic properties Anabaena Ferredoxin.

Analysis of the hyperfine-shifted nitrogen signals in the 15N NMR spectra of oxidized Anabaena 7120 heterocyst ferredoxin shows anomalous 15N T1 relaxation behavior in the nitrogens of the cysteinyl ligands to the cluster and suggests that electron-nuclear dipole-dipole interactions provide the dominant relaxation mechanism for the 15N nuclei. The simple dipolar model fails for short nitrogen distances from the cluster and this failure may be due to degradation of the point-dipole approximation and overlooking the delocalization of unpaired electron density from iron ions to other atoms.

A study of the diamagnetic proton resonance sequence-specific assignment of the Anabaena 7120 heterocyst ferredoxin revealed that heterocyst and vegetative ferredoxin differential specificity is due to local mobility and structural differences. Secondary structure features of the oxidized form included four beta-sheets and two alpha-helices.

Escherichia coli has been widely used as a host to produce recombinant proteins. However, because of its prokaryotic nature, it often fails to produce eukaryotic proteins in their native or correctly folded forms. To devise a systematic way to find such a condition that produces a large amount of correctly folded proteins, we chose to perturb the composition of intracellular metabolites by imposing different stresses such as the addition of sorbitol, NaCl, pH, and ethanol, and to monitor which metabolites showed the largest differences in the case of successful protein production. We profiled the metabolites under stresses by using two-dimensional NMR spectroscopy, and identified 39 metabolites after the protein production phase. We found that mild NaCl stress increased the yield of the target protein. Mildly high pH stress also helped in producing more of the target protein, and there is likely a connection between the isoelectric point of the target protein and the pH of medium for a higher yield. Exogenous sorbitol appeared to have been used to produce an osmoprotectant (trehalose) or direct energy source (glucose); thus it is advised that its use should not include isotopic labeling. Mild ethanol stress did not cause any significant change either in the metabolite profile or in the target protein production. We hope this work might shed light on how to improve production of the target protein by adjusting the external environment. [ABSTRACT FROM AUTHOR]

Selective isotope labeling was employed to analyze the origin of the paramagnetically shifed 1H, 2H and 15N resonances of T4MOC, a soluble Rieske ferredoxin component of toluene 4-monooxygenase. The paramagnetic H NMR signals from T4MOC differ from those of the plant-type and vertebrate ferredoxins, and assignments of these signals to residue and atom type in T4MOC question particular 1H NMR assignments made for the Xanthobacter Rieske center. The pattern of H-bonding to the iron-sulfur cluster of T4MOC is discussed.