88 research outputs found
Modulation of enrofloxacin binding in OmpF by Mg2+ as revealed by the analysis of fast flickering single-porin current
Get PDFOne major determinant of the efficacy of antibiotics on Gram-negative bacteria is the passage through the outer membrane. During transport of the fluoroquinolone enrofloxacin through the trimeric outer membrane protein OmpF of Escherichia coli, the antibiotic interacts with two binding sites within the pore, thus partially blocking the ionic current. The modulation of one affinity site by Mg2+ reveals further details of binding sites and binding kinetics. At positive membrane potentials, the slow blocking events induced by enrofloxacin in Mg2+-free media are converted to flickery sojourns at the highest apparent current level (all three pores flickering). This indicates weaker binding in the presence of Mg2+. Analysis of the resulting amplitude histograms with beta distributions revealed the rate constants of blocking (k(OB)) and unblocking (k(BO)) in the range of 1,000 to 120,000 s(-1). As expected for a bimolecular reaction, k(OB) was proportional to blocker concentration and k(BO) independent of it. k(OB) was approximately three times lower for enrofloxacin coming from the cis side than from the trans side. The block was not complete, leading to a residual conductivity of the blocked state being similar to 25% of that of the open state. Interpretation of the results has led to the following model: fast flickering as caused by interaction of Mg2+ and enrofloxacin is related to the binding site at the trans side, whereas the cis site mediates slow blocking events which are also found without Mg2+. The difference in the accessibility of the binding sites also explains the dependency of k(OB) on the side of enrofloxacin addition and yields a means of determining the most plausible orientation of OmpF in the bilayer. The voltage dependence suggests that the dipole of the antibiotic has to be adequately oriented to facilitate binding
Expression and regulation of Homer in human skeletal muscle during neuromuscular junction adaptation to disuse and exercise.
Get PDFProtein calcium sensors of the Homer family have been proposed to modulate the activity of various ion channels and nuclear factor of activated T cells (NFAT), the transcription factor modulating skeletal muscle differentiation. We monitored Homer expression and subcellular localization in human skeletal muscle biopsies following 60 d of bedrest [Second Berlin Bedrest Study (BBR2-2)]. Soleus (SOL) and vastus lateralis (VL) biopsies were taken at start (pre) and at end (end) of bedrest from healthy male volunteers of a control group without exercise (CTR; n=9), a resistive-only exercise group (RE; n=7), and a combined resistive/vibration exercise group (RVE; n=7). Confocal analysis showed Homer immunoreactivity at the postsynaptic microdomain of the neuromuscular junction (NMJ) at bedrest start. After bedrest, Homer immunoreactivity decreased (CTR), remained unchanged (RE), or increased (RVE) at the NMJ. Homer2 mRNA and protein were differently regulated in a muscle-specific way. Activated NFATc1 translocates from cytoplasm to nucleus; increased amounts of NFATc1-immunopositive slow-type myonuclei were found in RVE myofibers of both muscles. Pulldown assays identified NFATc1 and Homer as molecular partners in skeletal muscle. A direct motor nerve control of Homer2 was confirmed in rat NMJs by in vivo denervation. Homer2 is localized at the NMJ and is part of the calcineurin-NFATc1 signaling pathway. RVE has additional benefit over RE as countermeasure preventing disuse-induced neuromuscular maladaptation during bedrest
Structure-Activity Analysis of the Dermcidin-derived Peptide DCD-1L, an Anionic Antimicrobial Peptide Present in Human Sweat
No full textDermcidin encodes the anionic amphiphilic peptide DCD-1L, which displays a broad spectrum of antimicrobial activity under conditions resembling those in human sweat. Here, we have investigated its mode of antimicrobial activity. We found that DCD-1L interacts preferentially with negatively charged bacterial phospholipids with a helix axis that is aligned flat on a lipid bilayer surface. Upon interaction with lipid bilayers DCD-1L forms oligomeric complexes that are stabilized by Zn(2+). DCD-1L is able to form ion channels in the bacterial membrane, and we propose that Zn(2+)-induced self-assembly of DCD-1L upon interaction with bacterial lipid bilayers is a prerequisite for ion channel formation. These data allow us for the first time to propose a molecular model for the antimicrobial mechanism of a naturally processed human anionic peptide that is active under the harsh conditions present in human sweat
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Author Correction: Enhanced tenacity of mycobacterial aerosols from necrotic neutrophils
Get PDFThe original version of this Article contained errors within the affiliations section. Affiliation 4 was incorrectly given as ‘Leibniz Research Alliance INFECTIONS’21, Leipzig, Germany’. The correct affiliation is listed below: Leibniz Research Alliance INFECTIONS’21, Borstel, 23845, Germany Also, Affiliation 5 was incorrectly given as ‘German Center for Infection Research, TTU-TB, Borstel, 23845, Germany’. The correct affiliation is listed below: German Center for Infection Research (DZIF), Partner Site Hamburg-Lübeck-Borstel, Germany. Finally, the original HTML version of this Article omitted an affiliation for G. Gabriel. The correct affiliations for G. Gabriel are listed below: Heinrich Pette Institute, Leibniz Institute for Experimental Virology, Hamburg, 20251, Germany. Leibniz Research Alliance INFECTIONS’21, Borstel, 23845, Germany. German Center for Infection Research (DZIF), Partner Site Hamburg-Lübeck-Borstel, Germany. These errors have now been corrected in the PDF and HTML versions of the Article
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Crucial roles of charged saccharide moieties in survival of gram negative bacteria against protamine revealed by combination of grazing incidence x-ray structural characterizations and Monte Carlo simulations
Get PDFGrazing incidence x-ray scattering techniques and Monte Carlo (MC) simulations are combined to reveal the influence of molecular structure (genetic mutation) and divalent cations on the survival of gram negative bacteria against cationic peptides such as protamine. The former yields detailed structures of bacterial lipopolysaccharide (LPS) membranes with minimized radiation damages, while the minimal computer model based on the linearized Poisson-Boltzmann theory allows for the simulation of conformational changes of macromolecules (LPSs and peptides) that occur in the time scale of ms. The complementary combination of the structural characterizations and MC simulation demonstrates that the condensations of divalent ions (Ca2+ or Mg2+) in the negatively charged core saccharides are crucial for bacterial survival. © 2010 The American Physical Society.Fil: Oliveira, Rafael Gustavo. Technische Universitat München; Alemania. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones en Química Biológica de Córdoba. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Centro de Investigaciones en Química Biológica de Córdoba; ArgentinaFil: Schneck, Emanuel. Universität Heidelberg; Alemania. Technische Universitat München; AlemaniaFil: Quinn, Bonnie E.. St. Francis Xavier University; Canadá. Networks Of Centres Of Excellence; CanadáFil: Konovalov, Oleg V.. European Synchrotron Radiation; FranciaFil: Brandenburg, Klaus. Research Center Borstel; AlemaniaFil: Gutsmann, Thomas. Research Center Borstel; AlemaniaFil: Gill, Tom. Dalhousie University Halifax; Canadá. Networks Of Centres Of Excellence; CanadáFil: Hanna, Charles B.. Advanced Foods and Materials Network of Centres of Excellen; Canadá. Boise State University; AlemaniaFil: Pink, David A.. St. Francis Xavier University; Canadá. Networks Of Centres Of Excellence; CanadáFil: Tanaka, Motomu. Universität Heidelberg; Alemania. Networks Of Centres Of Excellence; Canadá. Technische Universitat München; Alemani
Sacrificial bonds and hidden length: unraveling molecular mesostructures in tough materials
No full textSacrificial bonds and hidden length in structural molecules and composites have been found to greatly increase the fracture toughness of biomaterials by providing a reversible, molecular-scale energy-dissipation mechanism. This mechanism relies on the energy, of order 100 eV, needed to reduce entropy and increase enthalpy as molecular segments are stretched after being released by the breaking of weak bonds, called sacrificial bonds. This energy is relatively large compared to the energy needed to break the polymer backbone, of order a few eV. In many biological cases, the breaking of sacrificial bonds has been found to be reversible, thereby additionally providing a "self-healing" property to the material. Due to the nanoscopic nature of this mechanism, single molecule force spectroscopy using an atomic force microscope has been a useful tool to investigate this mechanism. Especially when investigating natural molecular constructs, force versus distance curves quickly become very complicated. In this work we propose various types of sacrificial bonds, their combination, and how they appear in single molecule force spectroscopy measurements. We find that by close analysis of the force spectroscopy curves, additional information can be obtained about the molecules and their bonds to the native constructs
X-Ray Phase Contrast Imaging of Black Lipid Membranes: Studying membrane structure under native conditions
No full textStructural studies of Trehalose Dimycolate in model membrane systems: X-Ray Reflectometry and GISAXS experiments
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