Acceptor Subsite Mutants of NCC 2970 GtfB 4,3-α-Glucanotransferase Regulate the Ratio of (α1 → 3)/(α1 → 6) Linkages in Biosynthesized α-Glucans.

NCC 2970 GtfB (2970 GtfB) is the only characterized 4,3-α-glucanotransferase (4,3-α-GTase) in the glycoside hydrolase (GH) 70 family belonging to the GtfB subfamily. However, the mechanism for its (α1 → 3) linkage formation remains unclear, and the structural determinants of its linkage specificity remain to be explored. Here, sequence alignment and structural comparison were conducted to identify key amino acids that may be critical for linkage specificity.

Action and cooperation in alginate degradation by three enzymes from the human gut bacterium Bacteroides eggerthii DSM 20697.

Alginate is a polysaccharide consumed by humans in edible seaweed and different foods where it is applied as a texturizing hydrocolloid or in encapsulations of drugs and probiotics. While gut bacteria are found to utilize and ferment alginate to health-beneficial short-chain fatty acids, knowledge on the details of the molecular reactions is sparse. Alginates are composed of mannuronic acid (M) and its C-5 epimer guluronic acid (G).

Validation of smartwatch electrocardiogram intervals in children compared to standard 12 lead electrocardiograms.

Unlabelled: Lay people are now able to obtain one-lead electrocardiograms (ECG) using smartwatches, which facilitates documentation of arrhythmias. The accuracy of smartwatch derived ECG intervals has not been validated in children though. Home-based monitoring of ECG intervals using a smartwatch could improve monitoring of children, e.

Kinetics and mapping of Ca-driven calmodulin conformations on skeletal and cardiac muscle ryanodine receptors.

Calmodulin transduces [Ca] information regulating the rhythmic Ca cycling between the sarcoplasmic reticulum and cytoplasm during contraction and relaxation in cardiac and skeletal muscle. However, the structural dynamics by which calmodulin modulates the sarcoplasmic reticulum Ca release channel, the ryanodine receptor, at physiologically relevant [Ca] is unknown. Using fluorescence lifetime FRET, we resolve different structural states of calmodulin and Ca-driven shifts in the conformation of calmodulin bound to ryanodine receptor.

Tailor-Made α-Glucans by Engineering the Processivity of α-Glucanotransferases via Tunnel-Cleft Active Center Interconversions.

The function of polysaccharides is intimately associated with their size, which is largely determined by the processivity of transferases responsible for their synthesis. A tunnel active center architecture has been recognized as a key factor that governs processivity of several glycoside hydrolases (GHs), e.g.

Impact of Underweight, Overweight, and Obesity on Health-Related Quality of Life in Children with Tetralogy of Fallot Variants.

Traditional cardiovascular risk factors put patients with congenital heart disease (CHD) at increased risk for cardiovascular morbidity and mortality. The aim of this study was to evaluate whether body mass index (BMI) is associated with health-related quality of life (HRQoL) in patients with variants of Tetralogy of Fallot (TOF). Patients and parents of children with variants of TOF-CHD were asked to fill out the PedsQL 4.

N1019D Mutant of 121 4,6-α-Glucanotransferase GtfB Significantly Improved Catalytic Activity.

121 4,6-α-glucanotransferase GtfB ( 121 GtfB), belonging to glycoside hydrolase family 70 (GH70), synthesizes linear isomalto/malto polysaccharides having (α1→6) linkages attached to the nonreducing ends of (α1→4) linked maltose oligosaccharide segments using starch or maltodextrin as a substrate. Since 121 GtfB has low catalytic activity and efficiency, it leads to substrate regeneration and reduced substrate utilization. In this study, we superimposed the crystal structure of 121 GtfB-ΔNΔV with that of NCC 2613 GtfB-ΔNΔV ( 2613 GtfB-ΔNΔV) to identify the acceptor binding subsites +1 to +3 and constructed five single-residue mutants and a random mutagenesis of N1019.

Insights into the Structure-Function Relationship of GH70 GtfB α-Glucanotransferases from the Crystal Structure and Molecular Dynamic Simulation of a Newly Characterized N1 GtfB Enzyme.

α-Glucanotransferases of the CAZy family GH70 convert starch-derived donors to industrially important α-glucans. Here, we describe characteristics of a novel GtfB-type 4,6-α-glucanotransferase of high enzyme activity (60.8 U mg) from N1 (LrN1 GtfB), which produces surprisingly large quantities of soluble protein in heterologous expression (173 mg pure protein per L of culture) and synthesizes the reuteran-like α-glucan with (α1 → 6) linkages in linear chains and branch points.

Progress in cyclodextrins as important molecules regulating catalytic processes of glycoside hydrolases.

Cyclodextrins (CDs) are important starch derivatives and commonly comprise α-, β-, and γ-CDs. Their hydrophilic surface and hydrophobic inner cavity enable regulation of enzyme catalysis through direct or indirect interactions. Clarifying interactions between CDs and enzyme is of great value for enzyme screening, mechanism exploration, regulation of catalysis, and applications.

Bioinformatics and functional selection of GH77 4-α-glucanotransferases for potato starch modification.

4-α-glucanotransferases (4αGTs, EC 2.4.1.

Flagellar interference with plasmid uptake in biofilms: a joint experimental and modeling study.

Plasmid conjugation is a key facilitator of horizontal gene transfer (HGT), and plasmids encoding antibiotic resistance drive the increasing prevalence of antibiotic resistance. In natural, engineered, and clinical environments, bacteria often grow in protective biofilms. Therefore, a better understanding of plasmid transfer in biofilms is needed.

Functional Roles of N-Terminal Domains in Pullulanase from Human Gut .

Pullulanases are multidomain α-glucan debranching enzymes with one or more N-terminal domains (NTDs) including carbohydrate-binding modules (CBMs) and domains of unknown function (DUFs). To elucidate the roles of NTDs in NCFM pullulanase (Pul), two truncated variants, Δ41-Pul (lacking CBM41) and Δ(41+DUFs)-Pul (lacking CBM41 and two DUFs), were produced recombinantly. Pul recognized 1.

Sequential starch modification by branching enzyme and 4-α-glucanotransferase improves retention of curcumin in starch-alginate beads.

A new super-branched amylopectin with longer exterior chains was produced from normal maize starch by modification with branching enzyme followed by 4-α-glucanotransferase, and applied for co-entrapment of a curcumin-loaded emulsion in alginate beads. The network structure of the gel beads was obtained with Ca-cross-linked alginate and a modest load of retrograded starch. The dual enzyme modified starch contained more and longer α-1,6-linked branch chains than single enzyme modified and unmodified starches and showed superior resistance to digestive enzymes.

Ligand selectivity hotspots in serotonin GPCRs.

Serotonin is a neurotransmitter regulating numerous physiological processes also modulated by drugs, for example, schizophrenia, depression, migraine, and obesity. However, these drugs typically have adverse effects caused by promiscuous binding across 12 serotonin and more than 20 homologous receptors. Recently, structures of the entire serotonin receptor family uncovered molecular ligand recognition.

Three alginate lyases provide a new gut isolate with the ability to grow on alginate.

Humans consume alginate in the form of seaweed, food hydrocolloids, and encapsulations, making the digestion of this mannuronic acid (M) and guluronic acid (G) polymer of key interest for human health. To increase knowledge on alginate degradation in the gut, a gene catalog from human feces was mined for potential alginate lyases (ALs). The predicted ALs were present in nine species of the Bacteroidetes phylum, of which two required supplementation of an -acting AL, expected to mimic cross-feeding in the gut.

RyR2 Binding of an Antiarrhythmic Cyclic Depsipeptide Mapped Using Confocal Fluorescence Lifetime Detection of FRET.

Hyperactivity of cardiac sarcoplasmic reticulum (SR) ryanodine receptor (RyR2) Ca-release channels contributes to heart failure and arrhythmias. Reducing the RyR2 activity, particularly during cardiac relaxation (diastole), is a desirable therapeutic goal. We previously reported that the unnatural enantiomer () of an insect-RyR activator, verticilide, inhibits porcine and mouse RyR2 at diastolic (nanomolar) Ca and has in vivo efficacy against atrial and ventricular arrhythmia.

Sulfide in engineered methanogenic systems - Friend or foe?

Sulfide ions are regarded to be toxic to microorganisms in engineered methanogenic systems (EMS), where organic substances are anaerobically converted to products such as methane, hydrogen, alcohols, and carboxylic acids. A vast body of research has addressed solutions to mitigate process disturbances associated with high sulfide levels, yet the established paradigm has drawn the attention away from the multifaceted sulfide interactions with minerals, organics, microbial interfaces and their implications for performance of EMS. This brief review brings forward sulfide-derived pathways other than toxicity and with potential significance for anaerobic organic matter degradation.

Molecular Mechanism of a FRET Biosensor for the Cardiac Ryanodine Receptor Pathologically Leaky State.

Ca leak from cardiomyocyte sarcoplasmic reticulum (SR) via hyperactive resting cardiac ryanodine receptor channels (RyR2) is pro-arrhythmic. An exogenous peptide (DPc10) binding promotes leaky RyR2 in cardiomyocytes and reports on that endogenous state. Conversely, calmodulin (CaM) binding inhibits RyR2 leak and low CaM affinity is diagnostic of leaky RyR2.

The Effect of Low-Dose Glucocorticoids Over Two Years on Weight and Blood Pressure in Rheumatoid Arthritis: Individual Patient Data From Five Randomized Trials.

Background: Weight gain and hypertension are well known adverse effects of treatment with high-dose glucocorticoids.

Objective: To evaluate the effects of 2 years of low-dose glucocorticoid treatment in rheumatoid arthritis (RA).

Design: Pooled analysis of 5 randomized controlled trials with 2-year interventions allowing concomitant treatment with disease-modifying antirheumatic drugs.

Long-term fracture risk in rheumatoid arthritis: impact of early sustained DAS28-remission and restored function, progressive erosive disease, body mass index, autoantibody positivity and glucocorticoids. A cohort study over 10 years.

Background: Risk of fragility fractures in patients with rheumatoid arthritis (RA) is increased. Disease-related inflammation in RA is associated with low Bone Mineral Density (BMD). However, effects of specific disease factors on fracture occurrence and whether or not such disease effects are independent of BMD are unknown.

The Engineered Hexosaminidase OGA-D120N Is an Efficient -/-/-Glycoligase That Also Catalyzes Formation and Release of Oxazoline Donors for Cascade Syntheses with Glycosynthases or Transglycosylases.

Engineering glycoside hydrolases is a major route to obtaining catalysts forming glycosidic bonds. Glycosynthases, thioglycoligases, and transglycosylases represent the main strategies, each having advantages and drawbacks. Here, we show that an engineered enzyme from the GH84 family, the acid-base mutant OGA-D120N, is an efficient -, -, and -glycoligase, able to use , , , and nucleophiles.

Molecular Mechanism of a FRET Biosensor for the Cardiac Ryanodine Receptor Pathologically Leaky State.

Ca leak from cardiomyocyte sarcoplasmic reticulum (SR) via hyperactive resting cardiac ryanodine receptor channels (RyR2) is pro-arrhythmic. An exogenous peptide, (DPc10) detects leaky RyR2 in cardiomyocytes. Conversely, calmodulin (CaM) inhibits RyR2 leak.

A serologically assessed neo-epitope biomarker of cellular fibronectin degradation is related to pulmonary fibrosis.

Background: Idiopathic pulmonary fibrosis (IPF) is characterized by excessive extracellular matrix (ECM) remodeling, herein ECM degradation. Fibronectin (FN) is an important component of the ECM that is produced by multiple cell types, including fibroblasts. Extra domain B (EDB) is specific for a cellular FN isoform which is found in the ECM.