1,009 research outputs found
Genetic Characterization of Conserved Charged Residues in the Bacterial Flagellar Type III Export Protein FlhA
For assembly of the bacterial flagellum, most of flagellar proteins are transported to the distal end of the flagellum by the flagellar type III protein export apparatus powered by proton motive force (PMF) across the cytoplasmic membrane. FlhA is an integral membrane protein of the export apparatus and is involved in an early stage of the export process along with three soluble proteins, FliH, FliI, and FliJ, but the energy coupling mechanism remains unknown. Here, we carried out site-directed mutagenesis of eight, highly conserved charged residues in putative juxta- and trans-membrane helices of FlhA. Only Asp-208 was an essential acidic residue. Most of the FlhA substitutions were tolerated, but resulted in loss-of-function in the ΔfliH-fliI mutant background, even with the second-site flhB(P28T) mutation that increases the probability of flagellar protein export in the absence of FliH and FliI. The addition of FliH and FliI allowed the D45A, R85A, R94K and R270A mutant proteins to work even in the presence of the flhB(P28T) mutation. Suppressor analysis of a flhA(K203W) mutation showed an interaction between FlhA and FliR. Taken all together, we suggest that Asp-208 is directly involved in PMF-driven protein export and that the cooperative interactions of FlhA with FlhB, FliH, FliI, and FliR drive the translocation of export substrate
An energy transduction mechanism used in bacterial flagellar type III protein export
Flagellar proteins of bacteria are exported by a specific export apparatus. FliI ATPase forms a complex with FliH and FliJ and escorts export substrates from the cytoplasm to the export gate complex, which is made up of six membrane proteins. The export gate complex utilizes proton motive force across the cytoplasmic membrane for protein translocation, but the mechanism remains unknown. Here we show that the export gate complex by itself is a proton–protein antiporter that uses the two components of proton motive force, Δψ and ΔpH, for different steps of the protein export process. However, in the presence of FliH, FliI and FliJ, a specific binding of FliJ with an export gate membrane protein, FlhA, is brought about by the FliH–FliI complex, which turns the export gate into a highly efficient, Δψ-driven protein export apparatus
M153R Mutation in a pH-Sensitive Green Fluorescent Protein Stabilizes Its Fusion Proteins
BACKGROUND: Green fluorescent protein (GFP) and its fusion proteins have been used extensively to monitor and analyze a wide range of biological processes. However, proteolytic cleavage often removes GFP from its fusion proteins, not only causing a poor signal-to-noise ratio of the fluorescent images but also leading to wrong interpretations. METHODOLOGY/PRINCIPAL FINDINGS: Here, we report that the M153R mutation in a ratiometric pH-sensitive GFP, pHluorin, significantly stabilizes its fusion products while the mutant protein still retaining a marked pH dependence of 410/470 nm excitation ratio of fluorescence intensity. The M153R mutation increases the brightness in vivo but does not affect the 410/470-nm excitation ratios at various pH values. CONCLUSIONS/SIGNIFICANCE: Since the pHluorin(M153R) probe can be directly fused to the target proteins, we suggest that it will be a potentially powerful tool for the measurement of local pH in living cells as well as for the analysis of subcellular localization of target proteins
Distillation of Liquid Xenon to Remove Krypton
A high performance distillation system to remove krypton from xenon was
constructed, and a purity level of Kr/Xe = was
achieved. This development is crucial in facilitating high sensitivity low
background experiments such as the search for dark matter in the universe.Comment: 15 pages, 11 figure
Self-shielding effect of a single phase liquid xenon detector for direct dark matter search
Liquid xenon is a suitable material for a dark matter search. For future
large scale experiments, single phase detectors are attractive due to their
simple configuration and scalability. However, in order to reduce backgrounds,
they need to fully rely on liquid xenon's self-shielding property. A prototype
detector was developed at Kamioka Observatory to establish vertex and energy
reconstruction methods and to demonstrate the self-shielding power against
gamma rays from outside of the detector. Sufficient self-shielding power for
future experiments was obtained.Comment: 8 pages, 8 figure
Development of a low-background HPGe detector at Kamioka Observatory
A new ultra-low background high-purity germanium (HPGe) detector has been
installed at the Kamioka underground experimental site. The background count
rate in the energy range from 40 keV to 2700 keV is about 25% lower than that
of the first HPGe detector installed in 2016, which has the same detector
specification and similar shielding geometry. This paper describes the
shielding configuration, including the cleaning of the material surface, the
comparison of calibration data and simulation, the time variation of the
background spectra, the sample measurement procedure, and some results of the
radioactivity in the selected samples
Structural Insight into the Rotational Switching Mechanism of the Bacterial Flagellar Motor
Structural analysis of a clockwise-biased rotation mutant of the bacterial
flagellar rotor protein FliG provides a new model for the arrangement of FliG
subunits in the motor, and novel insights into rotation switching
Study of Non-Standard Neutrino Interactions with Atmospheric Neutrino Data in Super-Kamiokande I and II
In this paper we study non-standard neutrino interactions as an example of
physics beyond the standard model using atmospheric neutrino data collected
during the Super-Kamiokande I(1996-2001) and II(2003-2005) periods. We focus on
flavor-changing-neutral-currents (FCNC), which allow neutrino flavor
transitions via neutral current interactions, and effects which violate lepton
non-universality (NU) and give rise to different neutral-current
interaction-amplitudes for different neutrino flavors. We obtain a limit on the
FCNC coupling parameter, varepsilon_{mu tau}, |varepsilon_{mu tau}|<1.1 x
10^{-2} at 90%C.L. and various constraints on other FCNC parameters as a
function of the NU coupling, varepsilon_{e e}. We find no evidence of
non-standard neutrino interactions in the Super-Kamiokande atmospheric data.Comment: 12 Pages, 14 figures. To be submitted to Phys. Rev.
Search for Nucleon Decay into Charged Anti-lepton plus Meson in Super-Kamiokande I and II
Searches for a nucleon decay into a charged anti-lepton (e^+ or {\mu}^+) plus
a light meson ({\pi}^0, {\pi}^-, {\eta}, {\rho}^0, {\rho}^-, {\omega}) were
performed using the Super-Kamiokande I and II data. Twelve nucleon decay modes
were searched for. The total exposure is 140.9 kiloton \cdot years, which
includes a 91.7 kiloton \cdot year exposure (1489.2 live days) of
Super-Kamiokande-I and a 49.2 kiloton \cdot year exposure (798.6 live days) of
Super-Kamiokande-II. The number of candidate events in the data was consistent
with the atmospheric neutrino background expectation. No significant evidence
for a nucleon decay was observed in the data. Thus, lower limits on the nucleon
partial lifetime at 90% confidence level were obtained. The limits range from
3.6 \times 10^31 to 8.2 \times 10^33 years, depending on the decay modes.Comment: 25 pages, 18 figure
Baby MIND: A magnetised spectrometer for the WAGASCI experiment
The WAGASCI experiment being built at the J-PARC neutrino beam line will
measure the difference in cross sections from neutrinos interacting with a
water and scintillator targets, in order to constrain neutrino cross sections,
essential for the T2K neutrino oscillation measurements. A prototype Magnetised
Iron Neutrino Detector (MIND), called Baby MIND, is being constructed at CERN
to act as a magnetic spectrometer behind the main WAGASCI target to be able to
measure the charge and momentum of the outgoing muon from neutrino charged
current interactions.Comment: Poster presented at NuPhys2016 (London, 12-14 December 2016). Title +
4 pages, LaTeX, 6 figure
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