A systematic study of the influence of the packing density of proteins on their activity is performed with cytochrome c oxidase (CcO) from R. sphaeroides as an example. The protein was incorporated ...into a protein-tethered bilayer lipid membrane and CcO was genetically engineered with a histidine−tag, attached to Subunit II, and then tethered by an interaction with functionalized thiol compounds bound to a gold electrode. The packing density was varied by diluting the functionalized thiol with a nonfunctionalized thiol that does not bind to the enzyme. After attaching the CcO to the gold surface, a lipid bilayer was formed to incorporate the tethered proteins. The reconstituted protein−lipid bilayer was characterized by surface enhanced infrared reflection absorption spectroscopy (SEIRAS), electrochemical impedance spectroscopy, surface plasmon resonance, and atomic force microscopy. The activity of the proteins within the reconstituted bilayer was probed by direct electrochemical electron injection and was shown to be very sensitive to the packing density of protein molecules. At low surface density of CcO, the bilayer did not effectively form, and protein aggregates were observed, whereas at very high surface density, very little lipid is able to intrude between the closely packed proteins. In both of these cases, redox activity, measured by the efficiency to accept electrons, is low. Redox activity of the enzyme is preserved in the biomimetic structure but only at a moderate surface coverage in which a continuous lipid bilayer is present and the proteins are not forced to aggregate. Electrostatic and other interaction forces between protein molecules are held responsible for these effects.
Radiation hard ceramic RPC development Akindinov, A; Dreyer, J; Fan, X ...
Journal of physics. Conference series,
01/2017, Volume:
798, Issue:
1
Journal Article
Peer reviewed
Open access
We report recent advances in R&D on the Beam Fragmentation and T0 Counter (BFTC) for the CBM experiment, based on RPCs with floating electrodes made of resistive ceramic material. An optimal value of ...the ceramics bulk resistivity has been determined to be about 5·109 ·cm. RPCs with such electrodes show even characteristics and stable operation under particle fluxes of up to 150 kHz/cm2, with the detection efficiency above 90%.
In Germany, tuberculous lesions in slaughtered pigs due to infection with members of the
Mycobacterium avium complex are increasingly reported. Contaminated food originating from pig or other ...livestock is discussed as potential source of human infection.
M. avium isolates from man (
n
=
45), pig (
n
=
29), and cattle (
n
=
13) were characterised by restriction fragment length polymorphism (RFLP) with respect to insertion sequences IS
1245 and IS
901 as well as by
XbaI-based pulsed-field gel electrophoresis (PFGE) and the results were compared by computer cluster correlation analysis, to determine potential sources of infection in man. By PCR, 55% of animal isolates was identified as
M. avium subsp.
avium, and 45% as
M. a. hominissuis. All human isolates belonged to
M. a. hominissuis. IS
1245-RFLP and PFGE resulted in two distinct main groupings reflecting the two subspecies, and dividing the isolates into several subgroups. Animal isolates of
M. a. hominissuis were widely distributed within the subgroups of human isolates.
M. a. avium isolates, further discriminated by IS
901-RFLP, formed host-associated subgroups for animals. Comparison of RFLP patterns with those of PFGE resulted in different subgroups as well as different pairs of isolates with high similarities. Only two isolates exhibited identical patterns by both methods. In general, results of both methods support the possibility that
M. a. hominissuis isolates from livestock represent a source of infection for man, probably by common environmental reservoirs. There was no evidence of human infections caused by
M. a. avium in Germany.
In the last 20 years, 2-stage cryocoolers have been found to provide an optimum solution for a wide range of applications like low temperature physics, superconducting cold electronics, cryopumping ...and superconducting magnets. For a proper design of helium cryostats with significant cold masses connected to the first and second stages of cryocoolers, it is important to have a load map (also called "working field") in order to estimate cool-down and warm-up time periods. Such load maps are either not presented in the open literature sources for "high" temperature ranges, or just given by manufacturing companies for general information but without any guarantee. In the present paper, the load map of a 2-stage Sumitomo 415DP cryocooler in the wide temperature range of 40-400 K is presented.
The goal of health professional education programs is to produce competent graduates, with an ability to work collaboratively as effective healthcare team members. We explored the reflections of ...students and clinical facilitators, in response to participation in a structured interprofessional education (IPE) clinical placement program. In our qualitative study we used an exploratory case study design. In our analysis, we highlight the benefits of interprofessional practice. Key themes identified by students included: limited opportunities to engage in IPE across their course; lack of clarity around IPE; value of IPE for students, practitioners, and patient outcomes; and need for IPE opportunities to be integrated into placements. Key themes identified by the clinical facilitators included: being reminded of the value of IPE for students and patients; preparation for IPE placements need to be embedded in curricula; coordination and communication of IPE learning activities need to be clear for staff and students; and IPE should continue as part of the broader clinical education agenda. Our findings reinforce the notion that students and clinical facilitators value the importance of IPE for student learning within the clinical placement setting. The outcomes offer valuable insights for universities and hospital and health care contexts for setting up and implementing IPE activities, and we provide recommendations for improving ongoing IPE efforts within clinical placement setting.
Bioelectronic coupling of multi-redox-site membrane proteins was accomplished with cytochrome
c oxidase (C
cO) as an example. A biomimetic membrane system was used for the oriented immobilization of ...the C
cO oxidase on a metal electrode. When the protein is immobilized with the C
cO binding side directed toward the electrode and reconstituted in situ into a lipid bilayer, it is addressable by direct electron transfer to the redox centers. Electron transfer to the enzyme via the spacer, referred to as electronic wiring, shows an exceptionally high rate constant. This allows a kinetic analysis of all four consecutive electron transfer steps within the enzyme to be carried out. Electron transfer followed by rapid scan cyclic voltametry in combination with surface-enhanced resonance Raman spectroscopy provides mechanistic and structural information about the heme centers. Probing the enzyme under turnover conditions showed mechanistic insights into proton translocation coupled to electron transfer. This bioelectronic approach opens a new field of activity to investigate complex processes in a wide variety of membrane proteins.
Cytochrome
c oxidase (CcO) from
Paracoccus denitrificans was immobilized in a strict orientation via a his-tag attached to subunit I on a gold film and reconstituted in situ into a protein-tethered ...bilayer lipid membrane. In this orientation, the cytochrome
c (cyt
c) binding site is directed away from the electrode pointing to the outer side of the protein-tethered bilayer lipid membrane architecture. The CcO can thus be activated by cyt
c under aerobic conditions. Catalytic activity was monitored by impedance spectroscopy, as well as cyclic voltammetry. Cathodic and anodic currents of the CcO with cyt
c added to the bulk solution were shown to increase under aerobic compared to anaerobic conditions. Catalytic activity was considered in terms of repeated electrochemical oxidation/reduction of the CcO/cyt
c complex in the presence of oxygen. The communication of cyt
c bound to the CcO with the electrode is discussed in terms of a hopping mechanism through the redox sites of the enzyme. Simulations supporting this hypothesis are included.
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