1,359 research outputs found
Rational design of a (S)-selective-transaminase for asymmetric synthesis of (1S)-1-(1,1′-biphenyl-2-yl)ethanamine
Amine transaminases offer an environmentally sustainable synthesis route for the production of pure chiral amines. However, their catalytic efficiency toward bulky ketone substrates is greatly limited by steric hindrance and therefore presents a great challenge for industrial synthetic applications. We hereby report an example of rational transaminase enzyme design to help alleviate these challenges. Starting from the Vibrio fluvialis amine transaminase that has no detectable catalytic activity toward the bulky aromatic ketone 2-acetylbiphenyl, we employed a rational design strategy combining in silico and in vitro studies to engineer the transaminase enzyme with a minimal number of mutations, achieving an high catalytic activity and high enantioselectivity. We found that, by introducing two mutations W57G/R415A, detectable enzyme activity was achieved. The rationally designed variant, W57F/R88H/V153S/K163F/I259M/R415A/V422A, showed an improvement in reaction rate by more than 1716-fold toward the bulky ketone under study, producing the corresponding enantiomeric pure (S)-amine (enantiomeric excess (ee) value of >99%)
Molecular details of the unique mechanism of chloride transport by acyanobacterial rhodopsin
Microbial rhodopsins are well known as versatile and ubiquitous light-driven
ion transporters and photosensors. While the proton transport mechanism has
been studied in great detail, much less is known about various modes of anion
transport. Until recently, only two main groups of light-driven anion pumps
were known, archaeal halorhodopsins (HRs) and bacterial chloride pumps (known
as ClRs or NTQs). Last year, another group of cyanobacterial anion pumps with
a very distinct primary structure was reported. Here, we studied the chloride-
transporting photocycle of a representative of this new group,
Mastigocladopsis repens rhodopsin (MastR), using time-resolved spectroscopy in
the infrared and visible ranges and site-directed mutagenesis. We found that,
in accordance with its unique amino acid sequence containing many polar
residues in the transmembrane region of the protein, its photocycle features a
number of unusual molecular events not known for other anion-pumping
rhodopsins. It appears that light-driven chloride ion transfers by MastR are
coupled with translocation of protons and water molecules as well as
perturbation of several polar sidechains. Of particular interest is transient
deprotonation of Asp-85, homologous to the cytoplasmic proton donor of light-
driven proton pumps (such as Asp-96 of bacteriorhodopsin), which may serve as
a regulatory mechanism
Photochemical characterization of a novel fungal rhodopsin from Phaeosphaeria nodorum
Eukaryotic microbial rhodopsins are widespread bacteriorhodopsin-like proteins found in many lower eukaryotic groups including fungi. Many fungi contain multiple rhodopsins, some significantly diverged from the original bacteriorhodopsin template. Although few fungal rhodopsins have been studied biophysically, both fast-cycling light-driven proton pumps and slow-cycling photosensors have been found. The purpose of this study was to characterize photochemically a new subgroup of fungal rhodopsins, the so-called auxiliary group. The study used the two known rhodopsin genes from the fungal wheat pathogen, Phaeosphaeria nodorum. One of the genes is a member of the auxiliary group while the other is highly similar to previously characterized proton-pumping Leptosphaeria rhodopsin. Auxiliary rhodopsin genes from a range of species form a distinct group with a unique primary structure and are located in carotenoid biosynthesis gene cluster. Amino acid conservation pattern suggests that auxiliary rhodopsins retain the transmembrane core of bacteriorhodopsins, including all residues important for proton transport, but have unique polar intramembrane residues. Spectroscopic characterization of the two yeast-expressed Phaeosphaeria rhodopsins showed many similarities: absorption spectra, conformation of the retinal chromophore, fast photocycling, and carboxylic acid protonation changes. It is likely that both Phaeosphaeria rhodopsins are proton-pumping, at least in vitro.We suggest that auxiliary rhodopsins have separated from their ancestors fairly recently and have acquired the ability to interact with as yet unidentified transducers, performing a photosensory function without changing their spectral properties and basic photochemistry
Broadband velocity modulation spectroscopy of HfF^+: towards a measurement of the electron electric dipole moment
Precision spectroscopy of trapped HfF^+ will be used in a search for the
permanent electric dipole moment of the electron (eEDM). While this dipole
moment has yet to be observed, various extensions to the standard model of
particle physics (such as supersymmetry) predict values that are close to the
current limit. We present extensive survey spectroscopy of 19 bands covering
nearly 5000 cm^(-1) using both frequency-comb and single-frequency laser
velocity-modulation spectroscopy. We obtain high-precision rovibrational
constants for eight electronic states including those that will be necessary
for state preparation and readout in an actual eEDM experiment.Comment: 13 pages, 7 figures, 3 table
Recent observation of short range nucleon correlations in nuclei and their implications for the structure of nuclei and neutron stars
Novel processes probing the decay of nucleus after removal of a nucleon with
momentum larger than Fermi momentum by hard probes finally proved unambiguously
the evidence for long sought presence of short-range correlations (SRCs) in
nuclei. In combination with the analysis of large , A(e,e')X processes at
they allow us to conclude that (i) practically all nucleons with momenta
300 MeV/c belong to SRCs, consisting mostly of two nucleons, ii)
probability of such SRCs in medium and heavy nuclei is , iii) a fast
removal of such nucleon practically always leads to emission of correlated
nucleon with approximately opposite momentum, iv) proton removal from
two-nucleon SRCs in 90% of cases is accompanied by a removal of a neutron and
only in 10% by a removal of another proton. We explain that observed absolute
probabilities and the isospin structure of two nucleon SRCs confirm the
important role that tensor forces play in internucleon interactions. We find
also that the presence of SRCs requires modifications of the Landau Fermi
liquid approach to highly asymmetric nuclear matter and leads to a
significantly faster cooling of cold neutron stars with neutrino cooling
operational even for . The effect is even stronger for the
hyperon stars. Theoretical challenges raised by the discovered dominance of
nucleon degrees of freedom in SRCs and important role of the spontaneously
broken chiral symmetry in quantum chromodynamics (QCD) in resolving them are
considered. We also outline directions for future theoretical and experimental
studies of the physics relevant for SRCs.Comment: 74 pages. Review article, updated version to be published in
International Journal of Modern Physics
Hadronic properties of the S_{11}(1535) studied by electroproduction off the deuteron
Properties of excited baryonic states are investigated in the context of
electroproduction of baryon resonances off the deuteron. In particular, the
hadronic radii and the compositeness of baryon resonances are studied for
kinematic situations in which their hadronic reinteraction is the dominant
contribution. Specifically, we study the reaction at for kinematics in which the produced hadronic state reinteracts
predominantly with the spectator nucleon. A comparison of constituent quark
model and effective chiral Lagrangian calculations of the shows
substantial sensitivity to the structure of the produced resonance.Comment: 24 pages, 5 figure
Regulation of Progranulin Expression in Human Microglia and Proteolysis of Progranulin by Matrix Metalloproteinase-12 (MMP-12)
Background: The essential role of progranulin (PGRN) as a neurotrophic factor has been demonstrated by the discovery that haploinsufficiency due to GRN gene mutations causes frontotemporal lobar dementia. In addition to neurons, microglia in vivo express PGRN, but little is known about the regulation of PGRN expression by microglia. Goal: In the current study, we examined the regulation of expression and function of PGRN, its proteolytic enzyme macrophage elastase (MMP-12), as well as the inhibitor of PGRN proteolysis, secretory leukocyte protease inhibitor (SLPI), in human CNS cells. Methods: Cultures of primary human microglia and astrocytes were stimulated with the TLR ligands (LPS or poly IC), Th1 cytokines (IL-1/IFNc), or Th2 cytokines (IL-4, IL-13). Results were analyzed by Q-PCR, immunoblotting or ELISA. The roles of MMP-12 and SLPI in PGRN cleavage were also examined. Results: Unstimulated microglia produced nanogram levels of PGRN, and PGRN release from microglia was suppressed by the TLR ligands or IL-1/IFNc, but increased by IL-4 or IL-13. Unexpectedly, while astrocytes stimulated with proinflammatory factors released large amounts of SLPI, none were detected in microglial cultures. We also identified MMP-12 as a PGRN proteolytic enzyme, and SLPI as an inhibitor of MMP-12-induced PGRN proteolysis. Experiments employing PGRN siRNA demonstrated that microglial PGRN was involved in the cytokine and chemokine production following TLR3/4 activation
GWAS of Follicular Lymphoma Reveals Allelic Heterogeneity at 6p21.32 and Suggests Shared Genetic Susceptibility with Diffuse Large B-cell Lymphoma
Non-Hodgkin lymphoma (NHL) represents a diverse group of hematological
malignancies, of which follicular lymphoma (FL) is a prevalent subtype. A
previous genome-wide association study has established a marker, rs10484561 in
the human leukocyte antigen (HLA) class II region on 6p21.32 associated with
increased FL risk. Here, in a three-stage genome-wide association study,
starting with a genome-wide scan of 379 FL cases and 791 controls followed by
validation in 1,049 cases and 5,790 controls, we identified a second independent
FL–associated locus on 6p21.32, rs2647012
(ORcombined = 0.64,
Pcombined = 2×10−21)
located 962 bp away from rs10484561 (r2<0.1 in controls). After
mutual adjustment, the associations at the two SNPs remained genome-wide
significant (rs2647012:ORadjusted = 0.70,
Padjusted = 4×10−12;
rs10484561:ORadjusted = 1.64,
Padjusted = 5×10−15).
Haplotype and coalescence analyses indicated that rs2647012 arose on an
evolutionarily distinct haplotype from that of rs10484561 and tags a novel
allele with an opposite (protective) effect on FL risk. Moreover, in a follow-up
analysis of the top 6 FL–associated SNPs in 4,449 cases of other NHL
subtypes, rs10484561 was associated with risk of diffuse large B-cell lymphoma
(ORcombined = 1.36,
Pcombined = 1.4×10−7).
Our results reveal the presence of allelic heterogeneity within the HLA class II
region influencing FL susceptibility and indicate a possible shared genetic
etiology with diffuse large B-cell lymphoma. These findings suggest that the HLA
class II region plays a complex yet important role in NHL
Genetics of rheumatoid arthritis contributes to biology and drug discovery
A major challenge in human genetics is to devise a systematic strategy to integrate disease-associated variants with diverse genomic and biological datasets to provide insight into disease pathogenesis and guide drug discovery for complex traits such as rheumatoid arthritis (RA)1. Here, we performed a genome-wide association study (GWAS) meta-analysis in a total of >100,000 subjects of European and Asian ancestries (29,880 RA cases and 73,758 controls), by evaluating ~10 million single nucleotide polymorphisms (SNPs). We discovered 42 novel RA risk loci at a genome-wide level of significance, bringing the total to 1012–4. We devised an in-silico pipeline using established bioinformatics methods based on functional annotation5, cis-acting expression quantitative trait loci (cis-eQTL)6, and pathway analyses7–9 – as well as novel methods based on genetic overlap with human primary immunodeficiency (PID), hematological cancer somatic mutations and knock-out mouse phenotypes – to identify 98 biological candidate genes at these 101 risk loci. We demonstrate that these genes are the targets of approved therapies for RA, and further suggest that drugs approved for other indications may be repurposed for the treatment of RA. Together, this comprehensive genetic study sheds light on fundamental genes, pathways and cell types that contribute to RA pathogenesis, and provides empirical evidence that the genetics of RA can provide important information for drug discovery
A Cryogenic Silicon Interferometer for Gravitational-wave Detection
The detection of gravitational waves from compact binary mergers by LIGO has opened the era of gravitational wave astronomy, revealing a previously hidden side of the cosmos. To maximize the reach of the existing LIGO observatory facilities, we have designed a new instrument able to detect gravitational waves at distances 5 times further away than possible with Advanced LIGO, or at greater than 100 times the event rate. Observations with this new instrument will make possible dramatic steps toward understanding the physics of the nearby Universe, as well as observing the Universe out to cosmological distances by the detection of binary black hole coalescences. This article presents the instrument design and a quantitative analysis of the anticipated noise floor
- …