Pharmacogenetics Meets Metabolomics: Discovery of Tryptophan as a New Endogenous OCT2 Substrate Related to Metformin Disposition

Genetic polymorphisms of the organic cation transporter 2 (OCT2), encoded by SLC22A2, have been investigated in association with metformin disposition. A functional decrease in transport function has been shown to be associated with the OCT2 variants. Using metabolomics, our study aims at a comprehensive monitoring of primary metabolite changes in order to understand biochemical alteration associated with OCT2 polymorphisms and discovery of potential endogenous metabolites related to the genetic variation of OCT2. Using GC-TOF MS based metabolite profiling, clear clustering of samples was observed in Partial Least Square Discriminant Analysis, showing that metabolic profiles were linked to the genetic variants of OCT2. Tryptophan and uridine presented the most significant alteration in SLC22A2-808TT homozygous and the SLC22A2-808G>T heterozygous variants relative to the reference. Particularly tryptophan showed gene-dose effects of transporter activity according to OCT2 genotypes and the greatest linear association with the pharmacokinetic parameters (Clrenal, Clsec, Cl/F/kg, and Vd/F/kg) of metformin. An inhibition assay demonstrated the inhibitory effect of tryptophan on the uptake of 1-methyl-4-phenyl pyrinidium in a concentration dependent manner and subsequent uptake experiment revealed differential tryptophan-uptake rate in the oocytes expressing OCT2 reference and variant (808G>T). Our results collectively indicate tryptophan can serve as one of the endogenous substrate for the OCT2 as well as a biomarker candidate indicating the variability of the transport activity of OCT2

Time-to-Event Genome-Wide Association Study for Incident Cardiovascular Disease in People with Type 2 Diabetes Mellitus

BACKGROUND: Type 2 diabetes mellitus (T2D) confers a two- to three-fold increased risk of cardiovascular disease (CVD). However, the mechanisms underlying increased CVD risk among people with T2D are only partially understood. We hypothesized that a genetic association study among people with T2D at risk for developing incident cardiovascular complications could provide insights into molecular genetic aspects underlying CVD. METHODS: From 16 studies of the Cohorts for Heart & Aging Research in Genomic Epidemiology (CHARGE) Consortium, we conducted a multi-ancestry time-to-event genome-wide association study (GWAS) for incident CVD among people with T2D using Cox proportional hazards models. Incident CVD was defined based on a composite of coronary artery disease (CAD), stroke, and cardiovascular death that occurred at least one year after the diagnosis of T2D. Cohort-level estimated effect sizes were combined using inverse variance weighted fixed effects meta-analysis. We also tested 204 known CAD variants for association with incident CVD among patients with T2D. RESULTS: A total of 49,230 participants with T2D were included in the analyses (31,118 European ancestries and 18,112 non-European ancestries) which consisted of 8,956 incident CVD cases over a range of mean follow-up duration between 3.2 and 33.7 years (event rate 18.2%). We identified three novel, distinct genetic loci for incident CVD among individuals with T2D that reached the threshold for genome-wide significance ( P<5.0×10 -8): rs147138607 (intergenic variant between CACNA1E and ZNF648) with a hazard ratio (HR) 1.23, 95% confidence interval (CI) 1.15 - 1.32, P=3.6×10 -9, rs11444867 (intergenic variant near HS3ST1) with HR 1.89, 95% CI 1.52 - 2.35, P=9.9×10 -9, and rs335407 (intergenic variant between TFB1M and NOX3) HR 1.25, 95% CI 1.16 - 1.35, P=1.5×10 -8. Among 204 known CAD loci, 32 were associated with incident CVD in people with T2D with P<0.05, and 5 were significant after Bonferroni correction ( P<0.00024, 0.05/204). A polygenic score of these 204 variants was significantly associated with incident CVD with HR 1.14 (95% CI 1.12 - 1.16) per 1 standard deviation increase ( P=1.0×10 -16). CONCLUSIONS: The data point to novel and known genomic regions associated with incident CVD among individuals with T2D

Synchrotron serial crystallography at 11C beamline of Pohang Light Source-II

Serial crystallography (SX) makes a significant contribution for time-resolved studies and forms the base of structural analysis at room temperature, with minimal radiation damage. Even though X-ray free electron laser provides a femtosecond scale X-ray pulse, high accessibility of synchrotron facilities gives a merit for application of SX experiment. Therefore, we performed serial synchrotron crystallography (SSX) of lysozyme crystals at room-temperature. Both fixed target and injector-based methods were used for SX experiments to determine the structure of lysozyme crystals. Approximately 19,600 and 40,000 diffraction images were collected during 40 and 80 min, for fixed target and injectionbased methods, respectively in the SSX experiments. The 10 Hz synchrotron X-ray radiation of the 11C beamline of the Pohang accelerator laboratory was used and the crystal structures of lysozyme were determined at 1.89 Å and 1.80 Å resolutions, respectively. These results provide experimental clues for routine SX at room temperature in synchrotrons.22Ykciotherothe

Catalyzed Atomic Layer Deposition of Silicon Oxide at Ultralow Temperature Using Alkylamine

We report the catalyzed atomic layer deposition (ALD) of silicon oxide using Si₂Cl₆, H₂O, and various alkylamines. The density functional theory (DFT) calculations using the periodic slab model of the SiO₂ surface were performed for the selection of alternative Lewis base catalysts with high catalytic activities. During the first half-reaction, the catalysts with less steric hindrance such as pyridine would be more effective than bulky alkylamines despite lower nucleophilicity. On the other hand, during the second half-reaction, the catalysts with a high nucleophilicity such as triethylamine (Et₃N) would be more efficient because the steric hindrance is less critical. The in situ process monitoring shows that the calculated atomic charge is a good indicator for expecting the catalyst activity in the ALD reaction. The use of Et₃N in the second half-reaction was essential to improving the growth rate as well as the step coverage of the film because the Et₃N-catalyzed process deposited a SiO₂ film with a step coverage of 98% that is better than 93% of the pyridine-catalyzed process. The adsorption of pyridine, ammonia (NH₃), or trimethylamine (Me₃N) salts was more favorable than that of Et₃N, n-Pr₃N, or ⁱPr₃N salts. Therefore, Et₃N was expected to incorporate less amine salts in the film as compared to pyridine, and the compositional analyses confirmed that the concentrations of Cl and N by the Et₃N-catalyzed process were significantly lower than those by the pyridine-catalyzed process

BL-11C Micro-MX: a high-flux microfocus macromolecular-crystallography beamline for micrometre-sized protein crystals at Pohang Light Source II

BL-11C, a new protein crystallography beamline, is an in-vacuum undulator-based microfocus beamline used for macromolecular crystallography at the Pohang Accelerator Laboratory and it was made available to users in June 2017. The beamline is energy tunable in the range 5.0–20 keV to support conventional single- and multi-wavelength anomalous-dispersion experiments against a wide range of heavy metals. At the standard working energy of 12.659 keV, the monochromated beam is focused to 4.1 µm (V) × 8.5 µm (H) full width at half-maximum at the sample position and the measured photon flux is 1.3 × 1012 photons s−1. The experimental station is equipped with a Pilatus3 6M detector, a micro-diffractometer (MD2S) incorporating a multi-axis goniometer, and a robotic sample exchanger (CATS) with a dewar capacity of 90 samples. This beamline is suitable for structural determination of weakly diffracting crystalline substances, such as biomaterials, including protein, nucleic acids and their complexes. In addition, serial crystallography experiments for determining crystal structures at room temperature are possible. Herein, the current beamline characteristics, technical information for users and some recent scientific highlights are described.11Nsciescopu