Engineering of methionine-auxotroph via parallel evolution of two enzymes from direct-sulfurylation pathway enables its recovery in minimal medium.

Methionine biosynthesis relies on the sequential catalysis of multiple enzymes. , the main bacteria used in research and industry for protein production and engineering, utilizes the three-step trans-sulfurylation pathway catalyzed by L-homoserine O-succinyl transferase, cystathionine gamma synthase and cystathionine beta lyase to convert L-homoserine to L-homocysteine. However, most bacteria employ the two-step direct-sulfurylation pathway involving L-homoserine O-acetyltransferases and O-acetyl homoserine sulfhydrylase.

Conversion of methionine biosynthesis in Escherichia coli from trans- to direct-sulfurylation enhances extracellular methionine levels.

Methionine is an essential amino acid in mammals and a precursor for vital metabolites required for the survival of all organisms. Consequently, its inclusion is required in diverse applications, such as food, feed, and pharmaceuticals. Although amino acids and other metabolites are commonly produced through microbial fermentation, high-yield biosynthesis of L-methionine remains a significant challenge due to the strict cellular regulation of the biosynthesis pathway.

Plasma Cell Gingivitis: Clinical and Pathological Case Report.

Introduction: Plasma cell gingivitis (PCG) is a rare benign condition usually found on marginal and attached gingiva. This case details a generalized PCG, to include the management of the patient and clinicopathologic characteristics of the disorder.

Case Presentation: The patient, a 24-year-old African American female, was referred to the periodontics clinic for severe generalized gingival erythema and edema.

Piezoelectrically Tuned Multimode Cavity Search for Axion Dark Matter.

The μeV axion is a well-motivated extension to the standard model. The Axion Dark Matter eXperiment (ADMX) collaboration seeks to discover this particle by looking for the resonant conversion of dark-matter axions to microwave photons in a strong magnetic field. In this Letter, we report results from a pathfinder experiment, the ADMX "Sidecar," which is designed to pave the way for future, higher mass, searches.

Help-Seeking by Parental Caregivers of Individuals with Intellectual Disabilities and Dual Diagnosis.

This study examined the roles of stigma and of enabling factors in help-seeking by parental caregivers of individuals with intellectual disabilities and dual diagnosis. Questionnaires were completed by 195 family caregivers. Lower family stigma was related to higher personal enabling factors (e.

Search for Invisible Axion Dark Matter with the Axion Dark Matter Experiment.

This Letter reports the results from a haloscope search for dark matter axions with masses between 2.66 and 2.81  μeV.

aCORN: An experiment to measure the electron-antineutrino correlation coefficient in free neutron decay.

We describe an apparatus used to measure the electron-antineutrino angular correlation coefficient in free neutron decay. The apparatus employs a novel measurement technique in which the angular correlation is converted into a proton time-of-flight asymmetry that is counted directly, avoiding the need for proton spectroscopy. Details of the method, apparatus, detectors, data acquisition, and data reduction scheme are presented, along with a discussion of the important systematic effects.

The aCORN Backscatter-Suppressed Beta Spectrometer.

Backscatter of electrons from a beta spectrometer, with incomplete energy deposition, can lead to undesirable effects in many types of experiments. We present and discuss the design and operation of a backscatter-suppressed beta spectrometer that was developed as part of a program to measure the electronantineutrino correlation coefficient in neutron beta decay (aCORN). An array of backscatter veto detectors surrounds a plastic scintillator beta energy detector.

Cavity design for high-frequency axion dark matter detectors.

In an effort to extend the usefulness of microwave cavity detectors to higher axion masses, above ∼8 μeV (∼2 GHz), a numerical trade study of cavities was conducted to investigate the merit of using variable periodic post arrays and regulating vane designs for higher-frequency searches. The results show that both designs could be used to develop resonant cavities for high-mass axion searches. Multiple configurations of both methods obtained the scanning sensitivity equivalent to approximately 4 coherently coupled cavities with a single tuning rod.

Additive-mediated electrochemical synthesis of platelike copper crystals for methanol electrooxidation.

A room-temperature electrochemical approach to synthesizing anisotropic platelike copper microcrystals and nanocrystals in the presence of potassium bromide is presented. Morphological and elemental characterization was performed using SEM, TEM, and XRD to confirm the anisotropic morphology and crystal structure of the synthesized copper particles. A possible mechanism for explaining the anisotropic crystal growth is proposed on the basis of the preferential adsorption of bromide ions to selective crystal faces.

Disorders of the masticatory muscles.

Muscle disorders involving the masticatory muscles have been considered analogous to skeletal muscle disorders throughout the body. However, emerging research has shed new light on the varied etiology, clinical presentation, diagnosis, and treatment of myofascial pain and masticatory muscle disorders. This article reviews the etiology and classification of regional masticatory muscle disorders, the clinical examination of the patient, and evidence-based treatment recommendations.

Clinical assessment of patients with orofacial pain and temporomandibular disorders.

Accurate diagnosis of chronic pain disorders of the mouth, jaws, and face is frequently complex. It is common for patients with chronic orofacial pain to consult multiple clinicians and receive ineffective treatment before a correct diagnosis is reached. This problem is a significant public health concern.

Optimisation of methods for quantifying plasma mRNA levels from genes responsible for coronary artery plaque development and destabilization.

Aim: To investigate the hypothesis that in patients with coronary atherosclerosis it is possible to measure plasma mRNA levels from genes responsible for plaque development and destabilization.

Methods: Methods for RNA isolation, mRNA transcription and quantitative PCR were evaluated and optimised, in order to achieve reliable mRNA quantification. RESULTS mRNA level was possible to quantify from plasma of patients with coronary atherosclerosis, as well as from healthy volunteers, from genes encoding cathepsin S, cathepsin B, CD40 molecule, monocyte chemotactic protein 1, death-associated protein kinase 1, matrix metallopeptidase 9, vascular cell adhesion molecule 1 and phosphoglycerate kinase 1 (reference gene).

CTSS activation coexists with CD40 activation in human atheroma: evidence from plasma mRNA analysis.

Objectives: By the analysis of plasma mRNA levels, we tested the hypothesis that, in human atheroma, CTSS activation coexists with activation of CD40.

Design And Methods: mRNAs were isolated from plasma of patients with coronary atherosclerosis and quantified by real-time RT-PCR.

Results: CTSS mRNA levels correlated with CD40 mRNA levels, independently of observed traditional risk factors for atherosclerosis and pharmacological treatment.

Increased DAPK1 but decreased CCL2 plasma levels of nucleic acids in patients with stable angina.

Introduction: We hypothesized that patients with stable angina have increased plasma levels of mRNA from genes responsible for atherosclerotic plaque development and destabilisation, i.e. from death-associated protein kinase (DAPK1) and monocyte chemotactic protein-1 (CCL2).

Increased plasma levels of CATS mRNA but not CATB mRNA in patients with coronary atherosclerosis.

Background: We hypothesized that patients with coronary atherosclerosis have increased plasma levels of cathepsin S (CATS) and cathepsin B (CATB) mRNA, the genes that are involved in atherosclerotic plaque development and destabilization.

Methods: mRNAs were isolated from plasma of 67 patients with coronary atherosclerosis (29 with stable angina, 38 with acute coronary syndrome) and 33 healthy subjects as controls, transcribed to cDNA and quantified by real-time PCR.

Results: Plasma levels were successfully measured in all samples.

Proteolysis of prion protein by cathepsin S generates a soluble beta-structured intermediate oligomeric form, with potential implications for neurotoxic mechanisms.

Formation of PrP aggregates is considered to be a characteristic event in the pathogenesis of TSE diseases, accompanied by brain inflammation and neurodegeneration. Factors identified as contributing to aggregate formation are of interest as potential therapeutic targets. We report that in vitro proteolysis of ovine PrP(94-233) (at neutral pH and in the absence of denaturants) by the protease cathepsin S, a cellular enzyme that also shows enhanced expression in pathogenic conditions, occurs selectively in the region 135-156.

Anti-rheumatic gold compounds as sublethal modulators of monocytic LPS-induced cytokine secretion.

The objective of this study was to quantify the ability of sublethal concentrations of several gold compounds to differentially modulate the monocytic secretion of key cytokines that are important in the etiology of rheumatic diseases. Human THP1 monocytic cells were exposed to the anti-rheumatic drugs auranofin (AF), gold sodium thiomalate (GSTM) or HAuCl4 (Au(III)) for 24-72 h. Succinate dehydrogenase (SDH) activity of the monocytes was used to determine sublethal concentrations.

Crystal structure of NS-134 in complex with bovine cathepsin B: a two-headed epoxysuccinyl inhibitor extends along the entire active-site cleft.

The crystal structure of the inhibitor NS-134 in complex with bovine cathepsin B reveals that functional groups attached to both sides of the epoxysuccinyl reactive group bind to the part of active-site cleft as predicted. The -Leu-Pro-OH side binds to the primed binding sites interacting with the His110 and His111 residues with its C-terminal carboxy group, whereas the -Leu-Gly-Meu (-Leu-Gly-Gly-OMe) part (Meu, methoxycarbonylmethyl) binds along the non-primed binding sites. Comparison with the propeptide structures of cathepsins revealed that the binding of the latter part is least similar to the procathepsin B structure; this result, together with the two-residue shift in positioning of the Leu-Gly-Gly part, suggests that the propeptide structures of the cognate enzymes may not be the best starting point for the design of reverse binding inhibitors.

The role of cathepsin C in Papillon-Lefèvre syndrome, prepubertal periodontitis, and aggressive periodontitis.

We have previously reported that loss-of-function mutations in the cathepsin C gene (CTSC) result in Papillon-Lefèvre syndrome, an autosomal recessive condition characterized by palmoplantar keratosis and early-onset, severe periodontitis. Others have also reported CTSC mutations in patients with severe prepubertal periodontitis, but without any skin manifestations. The possible role of CTSC variants in more common types of non-mendelian, early-onset, severe periodontitis ("aggressive periodontitis") has not been investigated.

Structure of human dipeptidyl peptidase I (cathepsin C): exclusion domain added to an endopeptidase framework creates the machine for activation of granular serine proteases.

Dipeptidyl peptidase I (DPPI) or cathepsin C is the physiological activator of groups of serine proteases from immune and inflammatory cells vital for defense of an organism. The structure presented shows how an additional domain transforms the framework of a papain-like endopeptidase into a robust oligomeric protease-processing enzyme. The tetrahedral arrangement of the active sites exposed to solvent allows approach of proteins in their native state; the massive body of the exclusion domain fastened within the tetrahedral framework excludes approach of a polypeptide chain apart from its termini; and the carboxylic group of Asp1 positions the N-terminal amino group of the substrate.

[Fiber-enriched cereal for constipation and for hypercholesterolemia].

A mixed-grain, high-fiber cereal (Disivit) prepared from oats, corn, wheat and soybean was used to treat 20 patients with chronic constipation and 22 with hypercholesterolemia in double-blind, cross-over trials. Disivit (50 g/d, containing 12.5 g dietary fiber) was given to the constipated patients for 2 weeks and then a low-fiber placebo for another 2 weeks, and similarly for the hypercholesterolemic patients.