An enzymatic system for poly g-glutamate (PGA) synthesis in Bacillus subtilis, the PgsBCA system, was investigated. The gene-disruption experiment showed that the enzymatic system was the sole machinery of PGA synthesis in B. subtilis. We succeeded in achieving the enzymatic synthesis of elongated PGAs with the cell membrane of the Escherichia coli clone producing PgsBCA in the presence of ATP and D-glutamate. The enzyme preparation solubilized from the membrane with 8 mM Chaps catalyzed ADPforming ATP hydrolysis only in the presence of glutamate; the D-enantiomer was the best cosubstrate, followed by the L-enantiomer. Each component of the system, PgsB, PgsC, and PgsA, was translated in vitro and the glutamatedependent ATPase reaction was kinetically analyzed. The PGA synthetase complex, PgsBCA, was suggested to be an atypical amide ligase.

One-carbon metabolism in yeast is an essential process that relies on at least one of three one-carbon donor molecules: serine, glycine, or formate. By a combination of genetics and biochemistry we have shown how cells regulate the balance of one-carbon flow between the donors by regulating cytoplasmic serine hydroxymethyltransferase activity in a side reaction occurring in the presence of excess glycine. This control governs the level of 5,10-methylene tetrahydrofolate (5,10-CH 2 -H 4 folate) in the cytoplasm, which has a direct role in signaling transcriptional control of the expression of key genes, particularly those encoding the unique components of the glycine decarboxylase complex (GCV1, GCV2, and GCV3). Based on these and other observations, we propose a model for how cells balance the need to supplement their one-carbon pools when charged folates are limiting or when glycine is in excess. We also propose that under normal conditions, cytoplasmic 5,10-CH 2 -H 4 folate is mainly directed to generating methyl groups via methionine, whereas one-carbon units generated from glycine in mitochondria are more directed to purine biosynthesis. When glycine is in excess, 5,10-CH 2 -H 4 folate is decreased, and the regulation loop shifts the balance of generation of one-carbon units into the mitochondrion.

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Herein, we describe the fabrication of a magnetically retrievable nanocomposite adorned with highly active Pd nanoparticles (NPs) (MRN-Pd), which is useful for the efficient reduction of nitroaromatics in aqueous solution.

ImportanceIn patients with coronary artery disease, some guidelines recommend initial statin treatment with high-intensity statins to achieve at least a 50% reduction in low-density lipoprotein cholesterol (LDL-C). An alternative approach is to begin with moderate-intensity statins and titrate to a specific LDL-C goal. These alternatives have not been compared head-to-head in a clinical trial involving patients with known coronary artery disease.ObjectiveTo assess whether a treat-to-target strategy is noninferior to a strategy of high-intensity statins for long-term clinical outcomes in patients with coronary artery disease.Design, Setting, and ParticipantsA randomized, multicenter, noninferiority trial in patients with a coronary disease diagnosis treated at 12 centers in South Korea (enrollment: September 9, 2016, through November 27, 2019; final follow-up: October 26, 2022).InterventionsPatients were randomly assigned to receive either the LDL-C target strategy, with an LDL-C level between 50 and 70 mg/dL as the target, or high-intensity statin treatment, which consisted of rosuvastatin, 20 mg, or atorvastatin, 40 mg.Main Outcomes and MeasuresPrimary end point was a 3-year composite of death, myocardial infarction, stroke, or coronary revascularization with a noninferiority margin of 3.0 percentage points.ResultsAmong 4400 patients, 4341 patients (98.7%) completed the trial (mean [SD] age, 65.1 [9.9] years; 1228 females [27.9%]). In the treat-to-target group (n = 2200), which had 6449 person-years of follow-up, moderate-intensity and high-intensity dosing were used in 43% and 54%, respectively. The mean (SD) LDL-C level for 3 years was 69.1 (17.8) mg/dL in the treat-to-target group and 68.4 (20.1) mg/dL in the high-intensity statin group (n = 2200) (P = .21, compared with the treat-to-target group). The primary end point occurred in 177 patients (8.1%) in the treat-to-target group and 190 patients (8.7%) in the high-intensity statin group (absolute difference, –0.6 percentage points [upper boundary of the 1-sided 97.5% CI, 1.1 percentage points]; P < .001 for noninferiority).Conclusions and RelevanceAmong patients with coronary artery disease, a treat-to-target LDL-C strategy of 50 to 70 mg/dL as the goal was noninferior to a high-intensity statin therapy for the 3-year composite of death, myocardial infarction, stroke, or coronary revascularization. These findings provide additional evidence supporting the suitability of a treat-to-target strategy that may allow a tailored approach with consideration for individual variability in drug response to statin therapy.Trial RegistrationClinicalTrials.gov Identifier: NCT02579499

Glycine specifically induces genes encoding subunits of the glycine decarboxylase complex (GCV1, GCV2, and GCV3), and this is mediated by a fall in cytoplasmic levels of 5,10-methylenetetrahydrofolate caused by inhibition of cytoplasmic serine hydroxymethyltransferase. Here it is shown that this control system extends to genes for other enzymes of one-carbon metabolism and de novo purine biosynthesis. Northern analysis of the response to glycine demonstrated that the induction of the GCV genes and the induction of other amino acid metabolism genes are temporally distinct. The genomewide response to glycine revealed that several other genes are rapidly co-induced with the GCV genes, including SHM2, which encodes cytoplasmic serine hydroxymethyltransferase. These results were refined by examining transcript levels in an shm2⌬ strain (in which cytoplasmic 5,10-methylenetetrahydrofolate levels are reduced) and a met13⌬ strain, which lacks the main methylenetetrahydrofolate reductase activity of yeast and is effectively blocked at consumption of 5,10-methylene tetrahydrofolate for methionine synthesis. Glycine addition also caused a substantial transient disturbance to metabolism, including a sequence of changes in induction of amino acid biosynthesis and respiratory chain genes. Analysis of the glycine response in the shm2⌬ strain demonstrated that apart from the one-carbon regulon, most of these transient responses were not contingent on a disturbance to onecarbon metabolism. The one-carbon response is distinct from the Bas1p purine biosynthesis regulon and thus represents the first example of transcriptional regulation in response to activated one-carbon status.

This study sought to assess the antiviral efficacy of lamivudine (LMV) administered during third trimester to reduce maternal viraemia and to identify the emergence of LMV resistance. A prospective observational analysis was performed on 26 mothers with high viral load (>10⁷ IU/mL). Twenty-one women received LMV (treated group) for an average of 53 days (range 22-88 days), and the remaining five formed the untreated control group. Serum samples from two time points were used to measure HBV DNA levels and antiviral drug resistance. The LMV-treated women achieved a median HBV DNA reduction of 2.6-log10 IU/mL. Although end-of-treatment (EOT) HBV DNA in four (18%) LMV-treated women remained at >10(7) IU/mL (± 0.5 log IU/mL), no mother-to-baby transmission was observed. In contrast, a baby from the untreated mother was HBsAg positive at 9 months postpartum. Four technologies were used for drug resistance testing. Only ultra-deep pyrosequencing (UDPS) was sufficiently sensitive to detect minor viral variants down to <1%. UDPS showed that LMV therapy resulted in increased viral quasispecies diversity and positive selection of HBV variants with reverse transcriptase amino acid substitutions at sites associated with primary LMV resistance (rtM204I/V and rtA181T) in four (19%) women. These viral variants were detected mostly at low frequencies (0.63-5.92%) at EOT, but one LMV-treated mother had an rtA181T variant that increased from 2.2% pretherapy to 25.59% at EOT. This mother was also infected with the vaccine escape variant (sG145R), which was inhibited by LMV treatment. LMV therapy during late pregnancy only reduced maternal viraemia moderately, and drug-resistant viral variants emerged.