41 research outputs found
N-terminal lipid modification is required for the stable accumulation of CyanoQ in Synechocystis sp. PCC 6803
© 2016 Juneau et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. The CyanoQ protein has been demonstrated to be a component of cyanobacterial Photosystem II (PS II), but there exist a number of outstanding questions concerning its physical association with the complex. CyanoQ is a lipoprotein; upon cleavage of its transit peptide by Signal Peptidase II, which targets delivery of the mature protein to the thylakoid lumenal space, the N-terminal cysteinyl residue is lipid-modified. This modification appears to tether this otherwise soluble component to the thylakoid membrane. To probe the functional significance of the lipid anchor, mutants of the CyanoQ protein have been generated in Synechocystis sp. PCC 6803 to eliminate the N-terminal cysteinyl residue, preventing lipid modification. Substitution of the N-terminal cysteinyl residue with serine (Q-C22S) resulted in a decrease in the amount of detectable CyanoQ protein to 17% that of the wild-type protein. Moreover, the physical properties of the accumulated Q-C22S protein were consistent with altered processing of the CyanoQ precursor. The Q-C22S protein was shifted to a higher apparent molecular mass and partitioned in the hydrophobic phase in TX-114 phase-partitioning experiments. These results suggest that the hydrophobic N-terminal 22 amino acids were not properly cleaved by a signal peptidase. Substitution of the entire CyanoQ transit peptide with the transit peptide of the soluble lumenal protein PsbO yielded the Q-SS mutant and resulted in no detectable accumulation of the modified CyanoQ protein. Finally, the CyanoQ protein was present at normal amounts in the PS II mutant strains ÎpsbB and ÎpsbO, indicating that an association with PS II was not a prerequisite for stable CyanoQ accumulation. Together these results indicate that CyanoQ accumulation in Synechocystis sp. PCC 6803 depends on the presence of the N-terminal lipid anchor, but not on the association of CyanoQ with the PS II complex
CANDELS: The Cosmic Assembly Near-infrared Deep Extragalactic Legacy Survey
The Cosmic Assembly Near-infrared Deep Extragalactic Legacy Survey (CANDELS)
is designed to document the first third of galactic evolution, over the
approximate redshift (z) range 8--1.5. It will image >250,000 distant galaxies
using three separate cameras on the Hubble Space Telescope, from the
mid-ultraviolet to the near-infrared, and will find and measure Type Ia
supernovae at z>1.5 to test their accuracy as standardizable candles for
cosmology. Five premier multi-wavelength sky regions are selected, each with
extensive ancillary data. The use of five widely separated fields mitigates
cosmic variance and yields statistically robust and complete samples of
galaxies down to a stellar mass of 10^9 M_\odot to z \approx 2, reaching the
knee of the ultraviolet luminosity function (UVLF) of galaxies to z \approx 8.
The survey covers approximately 800 arcmin^2 and is divided into two parts. The
CANDELS/Deep survey (5\sigma\ point-source limit H=27.7 mag) covers \sim 125
arcmin^2 within GOODS-N and GOODS-S. The CANDELS/Wide survey includes GOODS and
three additional fields (EGS, COSMOS, and UDS) and covers the full area to a
5\sigma\ point-source limit of H \gtrsim 27.0 mag. Together with the Hubble
Ultra Deep Fields, the strategy creates a three-tiered "wedding cake" approach
that has proven efficient for extragalactic surveys. Data from the survey are
nonproprietary and are useful for a wide variety of science investigations. In
this paper, we describe the basic motivations for the survey, the CANDELS team
science goals and the resulting observational requirements, the field selection
and geometry, and the observing design. The Hubble data processing and products
are described in a companion paper.Comment: Submitted to Astrophysical Journal Supplement Series; Revised
version, subsequent to referee repor
Overview of the instrumentation for the Dark Energy Spectroscopic Instrument
The Dark Energy Spectroscopic Instrument (DESI) embarked on an ambitious 5 yr survey in 2021 May to explore the nature of dark energy with spectroscopic measurements of 40 million galaxies and quasars. DESI will determine precise redshifts and employ the baryon acoustic oscillation method to measure distances from the nearby universe to beyond redshift z > 3.5, and employ redshift space distortions to measure the growth of structure and probe potential modifications to general relativity. We describe the significant instrumentation we developed to conduct the DESI survey. This includes: a wide-field, 3.°2 diameter prime-focus corrector; a focal plane system with 5020 fiber positioners on the 0.812 m diameter, aspheric focal surface; 10 continuous, high-efficiency fiber cable bundles that connect the focal plane to the spectrographs; and 10 identical spectrographs. Each spectrograph employs a pair of dichroics to split the light into three channels that together record the light from 360â980 nm with a spectral resolution that ranges from 2000â5000. We describe the science requirements, their connection to the technical requirements, the management of the project, and interfaces between subsystems. DESI was installed at the 4 m Mayall Telescope at Kitt Peak National Observatory and has achieved all of its performance goals. Some performance highlights include an rms positioner accuracy of better than 0.âł1 and a median signal-to-noise ratio of 7 of the [O ii] doublet at 8 Ă 10â17 erg sâ1 cmâ2 in 1000 s for galaxies at z = 1.4â1.6. We conclude with additional highlights from the on-sky validation and commissioning, key successes, and lessons learned
PS II Protein Accumulation in other Genetic Backgrounds.
<p>Membranes from Lane 1, wild type; Lane 2, <i>ÎpsbO</i>; and, Lane 3, <i>ÎpsbB</i>, corresponding to 5 ÎŒg Chl per lane. After immunoblotting, specific antibodies against the CP47, CyanoQ and PsbU proteins were used to detect these components by chemiluminescence.</p
TX-114 Phase Partitioning.
<p>Thylakoid membranes from Q-Ctrl (A) and Q-C22S (B) were partitioned into an aqueous fraction and a TX-114 detergent fraction. The total membranes along with the aqueous and TX-114 phases were separated by LDS-PAGE. Proteins were detected using Coomassie blue. After immunoblotting, specific antibodies against the CP47, PsbO and CyanoQ proteins were used to detect these components by chemiluminescence.</p
<i>CyanoQ</i> Gene Structure and Mutant Constructs.
<p><i>CyanoQ</i> Gene Structure and Mutant Constructs.</p
Detection of CyanoQ in Membrane Samples.
<p>Membranes from Q-Ctrl were used as a standard series for CyanoQ detection. Lane 1, 5 ÎŒg Chl; Lane 2, 2.5 ÎŒg Chl; Lane 3, 1.25 ÎŒg Chl; Lane 4, 0.5 ÎŒg Chl; Lane 5 0.25 ÎŒg Chl, Lane 6, 0.05 ÎŒg Chl. Membranes from Q-C22S (Lane 7, 5 ÎŒg Chl; Lane 8, 10 ÎŒg Chl; Lane 9, 12.5 ÎŒg Chl) and Q-SS (Lane 10, 5 ÎŒg Chl; Lane 11, 10 ÎŒg Chl; Lane 12, 12.5 ÎŒg Chl). After immunoblotting all lanes were probed with an anti-CyanoQ protein antibody and detected by chemiluminescence.</p
N-Terminal Lipid Modification Is Required for the Stable Accumulation of CyanoQ in Synechocystis sp. PCC 6803
The CyanoQ protein has been demonstrated to be a component of cyanobacterial Photosystem II (PS II), but there exist a number of outstanding questions concerning its physical association with the complex. CyanoQ is a lipoprotein; upon cleavage of its transit peptide by Signal Peptidase II, which targets delivery of the mature protein to the thylakoid lumenal space, the N-terminal cysteinyl residue is lipid-modified. This modification appears to tether this otherwise soluble component to the thylakoid membrane. To probe the functional significance of the lipid anchor, mutants of the CyanoQ protein have been generated in Synechocystis sp. PCC 6803 to eliminate the N-terminal cysteinyl residue, preventing lipid modification. Substitution of the N-terminal cysteinyl residue with serine (Q-C22S) resulted in a decrease in the amount of detectable CyanoQ protein to 17% that of the wild-type protein. Moreover, the physical properties of the accumulated Q-C22S protein were consistent with altered processing of the CyanoQ precursor. The Q-C22S protein was shifted to a higher apparent molecular mass and partitioned in the hydrophobic phase in TX-114 phase-partitioning experiments. These results suggest that the hydrophobic N-terminal 22 amino acids were not properly cleaved by a signal peptidase. Substitution of the entire CyanoQ transit peptide with the transit peptide of the soluble lumenal protein PsbO yielded the Q-SS mutant and resulted in no detectable accumulation of the modified CyanoQ protein. Finally, the CyanoQ protein was present at normal amounts in the PS II mutant strains ÎpsbB and ÎpsbO, indicating that an association with PS II was not a prerequisite for stable CyanoQ accumulation. Together these results indicate that CyanoQ accumulation in Synechocystis sp. PCC 6803 depends on the presence of the N-terminal lipid anchor, but not on the association of CyanoQ with the PS II complex