Structural Evidence of a Major Conformational Change Triggered by Substrate Binding in DapE Enzymes: Impact on the Catalytic Mechanism

The X-ray crystal structure of the dapE-encoded N-succinyl-l,l-diaminopimelic acid desuccinylase from Haemophilus influenzae (HiDapE) bound by the products of hydrolysis, succinic acid and l,l-DAP, was determined at 1.95 Å. Surprisingly, the structure bound to the products revealed that HiDapE undergoes a significant conformational change in which the catalytic domain rotates ∼50° and shifts ∼10.1 Å (as measured at the position of the Zn atoms) relative to the dimerization domain. This heretofore unobserved closed conformation revealed significant movements within the catalytic domain compared to that of wild-type HiDapE, which results in effectively closing off access to the dinuclear Zn(II) active site with the succinate carboxylate moiety bridging the dinculear Zn(II) cluster in a μ-1,3 fashion forming a bis(μ-carboxylato)dizinc(II) core with a Zn–Zn distance of 3.8 Å. Surprisingly, His194.B, which is located on the dimerization domain of the opposing chain ∼10.1 Å from the dinuclear Zn(II) active site, forms a hydrogen bond (2.9 Å) with the oxygen atom of succinic acid bound to Zn2, forming an oxyanion hole. As the closed structure forms upon substrate binding, the movement of His194.B by more than ∼10 Å is critical, based on site-directed mutagenesis data, for activation of the scissile carbonyl carbon of the substrate for nucleophilic attack by a hydroxide nucleophile. Employing the HiDapE product-bound structure as the starting point, a reverse engineering approach called product-based transition-state modeling provided structural models for each major catalytic step. These data provide insight into the catalytic reaction mechanism and also the future design of new, potent inhibitors of DapE enzymes

Association between fertilin beta, protamines 1 and 2 and spermatid-specific linker histone H1-like protein mRNA levels, fertilization ability of human spermatozoa, and quality of preimplantation embryos.

Fertilization involves a series of cellular interactions culminating in the fusion of gamete membranes, creating a zygote and then an embryo. During the process of human fertilization in vivo or in conventional in vitro fertilization (IVF), sperm must be capable of undergoing the acrosome reaction, binding to the zona pellucida (ZP), and penetrating the ZP to fuse with the oolema. The key role in this process is played by fertilin beta. Protamines and histones are the proteins that bind to sperm chromatin and contribute in chromatin remodeling during early spermiogenesis. It has been suggested that these proteins may also participate in successful fertilization and embryo development. Using reverse transcription and real-time quantitative PCR reaction (QR-PCR) methods and zygote and embryo scoring, we compared fertilin beta, protamine 1 (PRM1), protamine 2 (PRM2), spermatid-specific linker histone 1 (HILS1) mRNAs levels, in vitro fertilization ability of mature spermatozoa, and quality of embryos obtained from in vitro fertilization (IVF). We found significantly lower contents of fertilin beta transcript in spermatozoa from patients in which IVF fertilization failed (

Evaluation of protamines 1 and 2 transcript contents in spermatozoa from asthenozoospermic men.

During mammalian spermatogenesis, the chromatin structure undergoes substantial condensation. The key role in this process is played by protamines 1 and 2 (PRM1, PRM2). We attempted to compare the levels of PRM1 and PRM2 transcripts in mature spermatozoa of normospermic and asthenozoospermic men. Human ejaculates from normozoospermic (n=70) and asthenozoospermic (n=100) donors were purified by centrifugation through discontinuous Percoll density gradient. RNA was isolated from spermatozoa according to the ChomczyĂąski and Sacchi method, treated with DNase I, and reverse-transcribed into cDNA. Using reverse transcription and real-time quantitative polymerase chain reaction analysis, we found a reduction in the levels of PRM1 and PRM2 transcripts in spermatozoa from asthenozoospermic men, as compared to controls (

The BSUIN project

Baltic Sea Underground Innovation Network (BSUIN) is an European Union funded project that extends capabilities of underground laboratories. The aim of the project is to join efforts in making the underground laboratories in the Baltic Sea Region’s more accessible for innovation, business development and science by improving the availability of information about the underground facilities, service offerings, user experience, safety and marketing.The development of standards for the characterization of underground laboratories will allow to compared them with each other. This will help you choose the best places for physical measurements such as neutrino physics or searching for dark matter. The project concerns laboratories where so far no measurements have been made, and even undergrounds where there are no organized laboratories yet.The description of the BSUIN project and the first results of characterization of natural radioactive background in underground laboratories will be presented ˙ The BSUIN Project is funded by Interreg Baltic Sea funding cooperation [2]

DM-like anomalies in neutron multiplicity spectra

Publisher Copyright: © 2022 Institute of Physics Publishing. All rights reserved.A new experiment collects data, since November 2019, at a depth of 210 m.w.e. in the Callio Lab in the Pyhasalmi mine in Finland. The setup, called NEMESIS (New Emma MEasurementS Including neutronS), incorporates infrastructure from the EMMA experiment with neutron and large-area plastic scintillator detectors. The experiment's primary aim is to combine muon tracking with position-sensitive neutron detection to measure precision yields, multiplicities, and lateral distributions of high-multiplicity neutron events induced by cosmic muons in various materials. The data are relevant for background evaluation of the deep-underground searches for Dark Matter (DM), neutrino-less double beta decay, etc. Preliminary analysis revealed anomalies in muon-suppressed neutron multiplicity spectra collected during a 344-day run (live time) with a 565 kg Pb target. The spectra, otherwise well described by an exponential fit, show three peaks at high multiplicities. Although still at a low statistical significance, these small excesses match the outcome of an earlier measurement. The nature of the anomalies remains unclear, but, in principle, they may be a signature of self-annihilation of a WIMP with a mass close to 13 GeV/c2. With that assumption, the expected cross-section would be around 10-42 cm2 for Spin-Dependent or 10-46 cm2 for Spin Independent interactions. We propose verifying this hypothesis with an upgraded NEMESIS experiment, able to collect an order of magnitude more data than this measurement. Based on the statistical uncertainty, analysis of the event rate indicates that cross-section limits for DM mass range of approximately 3-40 GeV/c2 can be investigated with such a setup.Peer reviewe

New NEMESIS Results

Funding Information: This work has been supported in part by the EU INTERREG for the Baltic Sea programme within the BSUIN project, and by the Polish Ministry of Science and Higher Education (Grant no. Funding Information: This work has been supported in part by the EU INTERREG for the Baltic Sea programme within the BSUIN project, and by the Polish Ministry of Science and Higher Education (Grant no. 3988/INTERREG BSR/2018/2). Publisher Copyright: © Copyright owned by the author(s) under the terms of the Creative Commons.Preliminary results from a 349-day run (live time) with a 565 kg Pb target and a 166-day background measurement are presented. Three minor anomalies were detected in muon-suppressed neutron multiplicity spectra. The multiplicities of these small excesses match the outcome of an earlier, similar but independent measurement. The nature of the anomalies remains unclear, but, in principle, they may be a signature of self-annihilation of a Weakly Interacting Massive Particle (WIMP) with a mass around 10 GeV/c2. If our interpretation is correct, the expected cross section would be of the order of 10-42 cm2 for Spin Dependent and 10-46 cm2 for Spin Independent interactions. Analysis of the event rate, based on the statistical uncertainty, indicates that cross-section limits for Dark Matter (DM) mass range of approximately 3-40 GeV/c2 can be investigated with an upgraded NEMESIS setup.Peer reviewe

NEMESIS setup for Indirect Detection of WIMPs

We summarize the evidence for DM-like anomalies in neutron multiplicity spectra collected underground with Pb targets by three independent experiments: NEMESIS (at 210 m.w.e.) NMDS (at 583 m.w.e.), and ZEPLIN-II (at 2850 m.w.e.). A new analysis shows small but persistent anomalies at high neutron multiplicities. Adjusted for differences in detection efficiencies, the positions of the anomalies are consistent between the three systems. Also, the intensities match when corrected for the acquisition time and estimated detection efficiency. While the three measurements are inconclusive when analyzed separately, together, they exclude a statistical fluke to better than one in a million. To prove the existence of the anomalies above the 5-sigma discovery threshold, we propose to upgrade the current NEMESIS setup. The upgrade concept and the critical components of the new experiment are described. The upgraded setup would already acquire the needed data sample during the first year of operation. Additional information, vital for the physics interpretation of the analysis, will be obtained with a Cu target.Peer reviewe

Spectroscopy of geo-neutrinos from 2056 days of Borexino data

We report an improved geo-neutrino measurement with Borexino from 2056 days of data taking. The present exposure is $(5.5\pm0.3)\times10^{31}$ proton$\times$yr. Assuming a chondritic Th/U mass ratio of 3.9, we obtain $23.7 ^{+6.5}_{-5.7} (stat) ^{+0.9}_{-0.6} (sys)$ geo-neutrino events. The null observation of geo-neutrinos with Borexino alone has a probability of $3.6 \times 10^{-9}$ (5.9$\sigma$). A geo-neutrino signal from the mantle is obtained at 98\% C.L. The radiogenic heat production for U and Th from the present best-fit result is restricted to the range 23-36 TW, taking into account the uncertainty on the distribution of heat producing elements inside the Earth.Comment: 4 pages, 4 figure

EBMT prospective observational study on allogeneic hematopoietic stem cell transplantation in T-prolymphocytic leukemia (T-PLL)

Preliminary data suggest that allogeneic stem cell transplantation (allo-SCT) may be effective in T-prolymphocytic leukemia (T-PLL). The purpose of the present observational study was to assess the outcome of allo-SCT in patients aged 65 years or younger with a centrally confirmed diagnosis of T-PLL. Patients were consecutively registered with the EBMT at the time of transplantation and followed by routine EBMT monitoring but with an extended dataset. Between 2007 and 2012, 37 evaluable patients (median age 56 years) were accrued. Pre-treatment contained alemtuzumab in 95% of patients. Sixty-two percent were in complete remission (CR) at the time of allo-SCT. Conditioning contained total body irradiation with 6 Gy or more (TBI6) in 30% of patients. With a median follow-up of 50 months, the 4-year non-relapse mortality, relapse incidence, progression-free (PFS) and overall survival were 32, 38, 30 and 42%, respectively. By univariate analysis, TBI6 in the conditioning was the only significant predictor for a low relapse risk, and an interval between diagnosis and allo-SCT of more than 12 months was associated with a lower NRM. This study confirms for the first time prospectively that allo-SCT can provide long-term disease control in a sizable albeit limited proportion of patients with T-PLL.Peer reviewe