Mobilization of Iron Stored in Bacterioferritin Is Required for Metabolic Homeostasis in Pseudomonas aeruginosa

Iron homeostasis offers a significant bacterial vulnerability because pathogens obtain essential iron from their mammalian hosts, but host-defenses maintain vanishingly low levels of free iron. Although pathogens have evolved mechanisms to procure host-iron, these depend on well-regulated iron homeostasis. To disrupt iron homeostasis, our work has targeted iron mobilization from the iron storage protein bacterioferritin (BfrB) by blocking a required interaction with its cognate ferredoxin partner (Bfd). The blockade of the BfrB–Bfd complex by deletion of the bfd gene (Δbfd) causes iron to irreversibly accumulate in BfrB. In this study we used mass spectrometry and NMR spectroscopy to compare the proteomic response and the levels of key intracellular metabolites between wild type (wt) and isogenic ΔbfdP. aeruginosa strains. We find that the irreversible accumulation of unusable iron in BfrB leads to acute intracellular iron limitation, even if the culture media is iron-sufficient. Importantly, the iron limitation and concomitant iron metabolism dysregulation trigger a cascade of events that lead to broader metabolic homeostasis disruption, which includes sulfur limitation, phenazine-mediated oxidative stress, suboptimal amino acid synthesis and altered carbon metabolism

Effect of cross polarization radiofrequency phases on signal phase

Utilizing phases of radio frequency (RF) pulses to manipulate spin dynamics is routine in NMR and MRI, leading to spectacular techniques like phase cycling. In a very different area, cross polarization (CP) also has a long history as part of a vast number of solid-state NMR pulse sequences. However, a detailed study devoted to the effect of CP RF phases on NMR signal, seems not to be readily available. From first principles, we arrive at a simple dependence of NMR signal on arbitrary CP RF phases, for static and MAS conditions, accompanied by experimental verification. In the process, the CP propagator emerges as a product of RF pulses and a period of free precession , conforming to coherence transfer pathway theory. The theoretical expressions may lend confidence for dealing with CP blocks with tunable phases in pulse sequences

Mobilization of Iron Stored in Bacterioferritin Is Required for Metabolic Homeostasis in Pseudomonas aeruginosa

Iron homeostasis offers a significant bacterial vulnerability because pathogens obtain essential iron from their mammalian hosts, but host-defenses maintain vanishingly low levels of free iron. Although pathogens have evolved mechanisms to procure host-iron, these depend on well-regulated iron homeostasis. To disrupt iron homeostasis, our work has targeted iron mobilization from the iron storage protein bacterioferritin (BfrB) by blocking a required interaction with its cognate ferredoxin partner (Bfd). The blockade of the BfrB–Bfd complex by deletion of the bfd gene (Δbfd) causes iron to irreversibly accumulate in BfrB. In this study we used mass spectrometry and NMR spectroscopy to compare the proteomic response and the levels of key intracellular metabolites between wild type (wt) and isogenic ΔbfdP. aeruginosa strains. We find that the irreversible accumulation of unusable iron in BfrB leads to acute intracellular iron limitation, even if the culture media is iron-sufficient. Importantly, the iron limitation and concomitant iron metabolism dysregulation trigger a cascade of events that lead to broader metabolic homeostasis disruption, which includes sulfur limitation, phenazine-mediated oxidative stress, suboptimal amino acid synthesis and altered carbon metabolism

Predicting Ion Association in Sodium Electrolytes: A Transferrable Model for Investigating Glymes

© 2017 American Chemical Society. Given the prior success in developing lithium batteries for similar purposes, many of the same types of solvent molecules and salt pairings have been investigated as electrolytes in sodium-ion and sodium-oxygen systems. Of these candidates, ether-containing electrolytes have emerged as a promising material as a result of their electrochemical stability and utility in tuning the pertinent electrochemistry. The ability for ethers to chelate metal ions provides a unique feature to ion solvation structure; however its role in changing the association of ions in solution has not been fully explored. By using computational simulations validated by FTIR and NMR spectroscopy, detailed descriptions of the changes to solvation structure as a result of chelation and concentration were investigated for a series of ethers (monoglyme to tetraglyme). From these simulations it can clearly be seen that with increasing chelation, ion association is diminished in a nonlinear fashion. For a monoglyme solvent, sodiums are entirely coordinated by triflates in solution, even at low concentrations. In contrast, tetraglymes retain a significant solvent separation of sodium cations from triflate anions even at high concentrations. The former implies that the utility of monoglyme and diglyme solvents for sodium-air batteries in specific is likely linked to their favoring ion association, while the poor performance of tetraglyme is a result of its excessive binding to sodium. Finally, triglyme was shown to produce anomalous behavior as a result of a mismatch between sodium coordination and steric interactions

Conformational Changes Associated with Post-Translational Modifications of Pro¹⁴³ in Skp1 of <i>Dictyostelium</i>A Dipeptide Model System

Prolyl hydroxylation and subsequent glycosylation of the E3^SCF ubiquitin ligase subunit Skp1 affects its conformation and its interaction with F-box proteins and, ultimately, O₂-sensing in the organism. Taking a reductionist approach to understand the molecular basis for these effects, a series of end-capped Thr-Pro dipeptides was synthesized, tracking the sequential post-translational modifications that occur in the protein. The conformation of the pyrrolidine ring in each compound was gauged via coupling constants (³<i>J</i>_{<i>H</i>α,<i>H</i>β}) and the electronegativity of the Cγ-substituents by chemical shifts (¹³C). The equilibrium between the <i>cis</i>–<i>trans</i> conformations about the central prolyl peptide bond was investigated by integration of signals corresponding to the two species in the ¹H NMR spectra over a range of temperatures. These studies revealed an increasing preference for the <i>trans</i>-conformation in the order Pro < Hyp < [α-(1,4)­GlcNAc]­Hyp. Rates for the forward and reverse reactions, determined by magnetization transfer experiments, demonstrated a reduced rate for the <i>trans</i>-to-<i>cis</i> conversion and a significant increase in the <i>cis</i>-to-<i>trans</i> conversion upon hydroxylation of the proline residue in the dipeptide. NOE experiments suggest that the Thr side chain pushes the sugar away from the pyrrolidine ring. These effects, which depended on the presence of the <i>N</i>-terminal Thr residue, offer a mechanism to explain altered properties of the corresponding full-length proteins