Biophysical Studies of the Membrane-Embedded and Cytoplasmic Forms of the Glucose-Specific Enzyme II of the E. coli Phosphotransferase System (PTS)

The glucose Enzyme II transporter complex of the Escherichia coli phosphotransferase system (PTS) exists in at least two physically distinct forms: a membrane-integrated dimeric form, and a cytoplasmic monomeric form, but little is known about the physical states of these enzyme forms. Six approaches were used to evaluate protein-protein and protein-lipid interactions in this system. Fluorescence energy transfer (FRET) using MBP-IIGlc-YFP and MBP-IIGlc-CFP revealed that the homodimeric Enzyme II complex in cell membranes is stable (FRET-) but can be dissociated and reassociated to the heterodimer only in the presence of Triton X100 (FRET+). The monomeric species could form a heterodimeric species (FRET+) by incubation and purification without detergent exposure. Formaldehyde cross linking studies, conducted both in vivo and in vitro, revealed that the dimeric MBP-IIGlc activity decreased dramatically with increasing formaldehyde concentrations due to both aggregation and activity loss, but that the monomeric MBP-IIGlc retained activity more effectively in response to the same formaldehyde treatments, and little or no aggregation was observed. Electron microscopy of MBP-IIGlc indicated that the dimeric form is larger than the monomeric form. Dynamic light scattering confirmed this conclusion and provided quantitation. NMR analyses provided strong evidence that the dimeric form is present primarily in a lipid bilayer while the monomeric form is present as micelles. Finally, lipid analyses of the different fractions revealed that the three lipid species (PE, PG and CL) are present in all fractions, but the monomeric micellar structure contains a higher percentage of anionic lipids (PG & CL) while the dimeric bilayer form has a higher percentage of zwitterion lipids (PE). Additionally, evidence for a minor dimeric micellar species, possibly an intermediate between the monomeric micellar and the dimeric bilayer forms, is presented. These results provide convincing evidence for interconvertible physical forms of Enzyme-IIGlc

Cryo-EM structure of the spinach cytochrome b6 f complex at 3.6 Å resolution.

The cytochrome b6 f (cytb6 f ) complex has a central role in oxygenic photosynthesis, linking electron transfer between photosystems I and II and converting solar energy into a transmembrane proton gradient for ATP synthesis1-3. Electron transfer within cytb6 f occurs via the quinol (Q) cycle, which catalyses the oxidation of plastoquinol (PQH2) and the reduction of both plastocyanin (PC) and plastoquinone (PQ) at two separate sites via electron bifurcation2. In higher plants, cytb6 f also acts as a redox-sensing hub, pivotal to the regulation of light harvesting and cyclic electron transfer that protect against metabolic and environmental stresses3. Here we present a 3.6 Å resolution cryo-electron microscopy (cryo-EM) structure of the dimeric cytb6 f complex from spinach, which reveals the structural basis for operation of the Q cycle and its redox-sensing function. The complex contains up to three natively bound PQ molecules. The first, PQ1, is located in one cytb6 f monomer near the PQ oxidation site (Qp) adjacent to haem bp and chlorophyll a. Two conformations of the chlorophyll a phytyl tail were resolved, one that prevents access to the Qp site and another that permits it, supporting a gating function for the chlorophyll a involved in redox sensing. PQ2 straddles the intermonomer cavity, partially obstructing the PQ reduction site (Qn) on the PQ1 side and committing the electron transfer network to turnover at the occupied Qn site in the neighbouring monomer. A conformational switch involving the haem cn propionate promotes two-electron, two-proton reduction at the Qn site and avoids formation of the reactive intermediate semiquinone. The location of a tentatively assigned third PQ molecule is consistent with a transition between the Qp and Qn sites in opposite monomers during the Q cycle. The spinach cytb6 f structure therefore provides new insights into how the complex fulfils its catalytic and regulatory roles in photosynthesis

Factors accounting for the association between anxiety and depression, and eczema: the Hordaland health study (HUSK)

<p>Abstract</p> <p>Background</p> <p>The association between anxiety and depression, and eczema is well known in the literature, but factors underlying this association remain unclear. Low levels of omega-3 fatty acids and female gender have been found to be associated with both depression and eczema. Somatization and health anxiety are known to be associated with anxiety and depression, further, somatization symptoms and health anxiety have also been found in several dermatological conditions. Accordingly, omega-3 fatty acid supplement, female gender, somatization and health anxiety are possible contributing factors in the association between anxiety and depression, and eczema. The aim of the study is to examine the relevance of proposed contributing factors for the association between anxiety and depression, and eczema, including, omega-3 fatty acid supplement, female gender, health anxiety and somatization.</p> <p>Methods</p> <p>Anxiety and depression was measured in the general population (n = 15715) employing the Hospital Anxiety and Depression Scale (HADS). Information on eczema, female gender, omega-3 fatty acid supplement, health anxiety and somatization was obtained by self-report.</p> <p>Results</p> <p>Somatization and health anxiety accounted for more than half of the association between anxiety/depression, and eczema, while the other factors examined were of minor relevance for the association of interest.</p> <p>Conclusions</p> <p>We found no support for female gender and omega-3 fatty acid supplement as contributing factors in the association between anxiety/depression, and eczema. Somatization and health anxiety accounted for about half of the association between anxiety/depression, and eczema, somatization contributed most. The association between anxiety/depression, and eczema was insignificant after adjustment for somatization and health anxiety. Biological mechanisms underlying the mediating effect of somatization are yet to be revealed.</p

Synthesis and characterization of a phosphaalkyne-bridged pentairon carbonyl cluster - crystal and molecular-structure of [(fe3se2(co)(8))(mu-pcbu(t))(fe2se(co)(6))]

The new cluster [{Fe3Se2(CO)8(})(mu-PCBu(t)){Fe2Se(CO)(6)}] has been isolated in moderate yield from the photolytic reaction of [Fe-2(CO)(6)(mu-Se-2] and the phosphaalkyne P=CBUt. Its structure has been established by single crystal X-ray diffraction methods: monoclinic, P2(1)/c (No. 14), a = 9.378(1) Angstrom, b = 17.605(3) Angstrom, c = 18.631(2) Angstrom, beta = 93.00(1)degrees, V = 3071.7 Angstrom(3) Z = 4, R = 7.6%, R' = 6.5%. The structure consists of a [Fe3Se2(CO)(8)] unit and a [Fe2Se(CO)(6)(PCBu(t))] unit linked by the phosphorus lone pair electrons of the phosphaalkyne

NOVEL ALKYNE AND PHOSPHAALKYNE COUPLING ON AN IR-4 CLUSTER - SYNTHESIS AND MOLECULAR-STRUCTURE OF [IR-4(MU-CO)(CO)(7)(MU(4)-ETA(3)-PH(2)PC(H)C(PH)PCBU(T))(MU-PPH(2))]

The cluster compound [(mu-H)Ir-4(CO)g(Ph(2)PCCPh)(mu-PPh(2))] 1 reacts with the phosphaalkyne Bu(t)CP to yield [Ir-4(mu-Co)(Co)(7){mu(4)-eta(3)-Ph(2)PC(H)C(Ph)PCBu(t)}(mu-PPh(2))] 3, containing the novel 2-phosphabutadienylphosphine fragment as a result of the coupling of Bu(t)CP with the diphenylphosphinoalkyne ligand and incorporation of the cluster bound H atom.161869187

First evidence for two different mu-eta(1)-eta(1)- and mu-eta(1)-eta(2)-co-ordination modes of the P3C2Bu2t ring of [Fe(eta(5)-P3C2Bu2t)(eta(5)-C5H5)] to a same cluster fragment: Synthesis and characterisation of [Ir-4(CO)(10){Fe(eta(5)-P3C2Bu2t)(eta(5)-C5H5)}] and X-ray molecular structure of the mu-eta(1)-eta(2)-isomer

Investigation of the solution structures of [Ir-4(CO)(11)L] (L = [Fe((eta(5)-P3C2Bu2t)(eta(5)-C5H5)] (1) and [Fe(eta(5)-P3C2Bu2t)(eta(5)-P2C3Bu3t)] (2) by C-13 and P-31 NMR spectroscopy showed that, at 163K, 1 exists in the form of two isomers with bridged and non-bridged structures, in a 1:0.15 ratio, respectively, whereas 2 exists only in the bridged form. At RT, 1,2 shift of the eta(5)-P3C2Bu2t ring was only observed for compound 2. Where as 2 loses CO readily in solution to give [Ir-4(CO)(10){mu-eta(1)-eta(1)-[Fe(eta(5)-P3C2Bu2t)(eta(5)-P2C3Bu3t)}] (3), activation with Me3NO was necessary to produce [Ir-4(CO)(10){[Fe(eta(5)-P3C2Bu2t)(eta(5)-C5H5)}] (4), obtained in the form of two non-interconverting isomers 4a and 4b, which were not able to be separated. A single crystal X-ray diffraction study of isomer 4a established that the [Fe(eta(5)-P3C2Bu2t)(eta(5)-C5H5)] ligand bridges one of the edges of the Ir4 tetrahedron, interacting via the lone electron pair of one of the adjacent P atoms and in an eta(2)- mode via the P-P double bond of the eta(5)-P3C2Bu2t ring and that all CO ligands are terminally bonded. Variable temperature P-31{H-1} NMR spectroscopy evidenced a fluxional process involving interactions between the Ir-1 and Ir-2 atoms and the lone pair on P-1, the P-1-P-2 bond, and the lone pair on P-2. According to multinuclear NMR, cluster 4b has similar structure to compound 3, with the eta(5)-P3C2Bu2t ring coordinated in a eta(1)-eta(1)-mode via the two adjacent P atoms, and all CO ligands bonded terminally.11440541