Journal: bioRxiv

Article Title: Mechanism of the secretion of the lanthipeptide nisin

doi: 10.1101/839423

Figure Lengend Snippet: ( A ) The supernatants of pre-NisA secreting L. lactis NZ9000 strains was analyzed by RP-HPLC and the amount of pre-NisA was determined. Amounts of secreted pre-peptides (nmol) are plotted against time (min) and the resulting curves were fitted by an allosteric sigmoidal fit. Modified NisA (mNisA, red) was secreted by strain NZ9000BTC (red dots) and can be preclude by nisT deletion (strain NZ9000BC, clear dot) or an ATP-deficient mutant (NZ9000BT H551A C, red rhomb). Dehydrated NisA (dNisA, blue) was secreted by strains NZ9000BTC H331A (blue rhomb) and NZ9000BT (blue dot), whereas unmodified NisA (uNisA, grey) was secreted by the strains NZ9000T (grey dots) and NZ9000TC (grey square). Dashed square shows a zoom-in on strains with lower secretion level. ( B ) The kinetic parameter of V max (nmol) of secreted peptides was plotted as bars against the various secretion systems. ( C ) The secretion rate of NisA molecules per NisT molecules was plotted against time (min) and fitted by linear regression. The slope represented the secretion rate of NisA•NisT −1 •min −1 for the strains NZ9000BTC and NZ9000T. All data represent secretion experiments from at least five different transformants and are represented as means ± SD (n=5). ++: WT secretion; o: low secretion; −: no secretion

Article Snippet: The culture supernatants containing pre-NisA variants were analyzed by RP-HPLC (Agilent Technologies 1260 Infinity II).

Techniques: Modification, Mutagenesis

Journal: bioRxiv

Article Title: Mechanism of the secretion of the lanthipeptide nisin

doi: 10.1101/839423

Figure Lengend Snippet: The supernatants of pre-NisA secreting L. lactis NZ9000 strains were employed for an RP-HPLC analysis. The pre-NisA variants (mNisA, dNisA and uNisA) were separated from other peptides in the supernatant by an acetonitrile/water gradient on a C-18 RP-HPLC column (left panel). The elution fractions (dashed square; middle panel) were further analyzed by MALDI-TOF-MS (right panel) to verify the correct mass. Integration of the corresponding peaks enables the determination of peptide amounts (nmol). Supernatant analysis of L. lactis strains ( A ) NZ9000BTC, ( B ) NZ9000BTC H331A , ( C ) NZ9000BT H551A C, ( D ) NZ9000BT, ( E ) NZ9000T, ( F ) NZ9000TC and ( G ) NZ9000BC.

Article Snippet: The culture supernatants containing pre-NisA variants were analyzed by RP-HPLC (Agilent Technologies 1260 Infinity II).

Techniques:

Journal: Frontiers in Pharmacology

Article Title: Binase Immobilized on Halloysite Nanotubes Exerts Enhanced Cytotoxicity toward Human Colon Adenocarcinoma Cells

doi: 10.3389/fphar.2017.00631

Figure Lengend Snippet: The scheme of the experimental work. (A) selection of conditions for loading of antitumor binase on halloysite nanotubes (HNTs) using solutions with different pH (NaAc, 10 mM Na-acetate buffer, pH 5.0; H 2 O, water of Milli-Q grade, pH 5.5; NaP, 10 mM Na-phosphate buffer, pH, 7.0; Tris-HCl, 0.25M Tris-HCl buffer pH 8.5); (B) optimal binase release from HNTs at pH 7.0. Due to the absence of difference in the quantity of released enzyme between sonicated and unsonicated samples the sonication step was excluded; (C) treatment of human colon adenocarcinoma cells Colo 320 with binase-loaded HNTs.

Article Snippet: The zeta-potential of binase-loaded HNTs was monitored using a Zetasizer Nano ZS instrument (Malvern, United Kingdom).

Techniques: Selection, Sonication

Journal: Frontiers in Pharmacology

Article Title: Binase Immobilized on Halloysite Nanotubes Exerts Enhanced Cytotoxicity toward Human Colon Adenocarcinoma Cells

doi: 10.3389/fphar.2017.00631

Figure Lengend Snippet: Characterization of binase loading on and release from HNTs. (A) The quantity of binase loaded onto HNTs at various pH (NaAc, 10 mM Na-acetate buffer, pH 5.0; H 2 O, water of Milli-Q grade, pH 5.5; NaP, 10 mM Na-phosphate buffer, pH, 7.0; Tris-HCl, 0.25M Tris-HCl buffer pH 8.5). (B) Zeta-potential of HNTs, binase-loaded HNTs and binase-loaded HNTs coated with dextrin. (C) Time-dependent release of binase from HNTs at neutral pH with and without sonication. Initial enzyme concentration in solutions was 1 mg/ml. The catalytic activity of binase in each solution is taken for 100%. Data represent mean ± SEM of three independent experiments; ∗ P < 0.05, ∗∗ P < 0.01 vs. control, ns, non-significant.

Article Snippet: The zeta-potential of binase-loaded HNTs was monitored using a Zetasizer Nano ZS instrument (Malvern, United Kingdom).

Techniques: Zeta Potential Analyzer, Sonication, Concentration Assay, Activity Assay, Control

Journal: Frontiers in Pharmacology

Article Title: Binase Immobilized on Halloysite Nanotubes Exerts Enhanced Cytotoxicity toward Human Colon Adenocarcinoma Cells

doi: 10.3389/fphar.2017.00631

Figure Lengend Snippet: Electron microscopy images of HNTs. SEM image of pure HNTs (A) , TEM image of pristine halloysite (B) and binase-loaded HNTs (C) . Arrows indicate the lumen of pristine nanotubes (B) and nanotubes loaded with binase (C) . Scale bar in (A) is 1 μm, in (B,C) is 100 nm.

Article Snippet: The zeta-potential of binase-loaded HNTs was monitored using a Zetasizer Nano ZS instrument (Malvern, United Kingdom).

Techniques: Electron Microscopy

Journal: Frontiers in Pharmacology

Article Title: Binase Immobilized on Halloysite Nanotubes Exerts Enhanced Cytotoxicity toward Human Colon Adenocarcinoma Cells

doi: 10.3389/fphar.2017.00631

Figure Lengend Snippet: Cytotoxicity of binase-loaded HNTs toward human colon adenocarcinoma Colo320 cells. Viability of cells in the absence of studied agents was taken as 100%. Data represent mean ± SEM of three independent experiments; ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗∗ P < 0.0001 vs. control, ns, non-significant.

Article Snippet: The zeta-potential of binase-loaded HNTs was monitored using a Zetasizer Nano ZS instrument (Malvern, United Kingdom).

Techniques: Control

Journal: Frontiers in Pharmacology

Article Title: Binase Immobilized on Halloysite Nanotubes Exerts Enhanced Cytotoxicity toward Human Colon Adenocarcinoma Cells

doi: 10.3389/fphar.2017.00631

Figure Lengend Snippet: Fluorescence microscopy of Colo 320 cells after 24 h of incubation (A) and simultaneous treatment by binase (B) , HNTs (C) , and HNTs loaded with binase (D) . Cells were stained with 3,3′-dihexyloxacarbocyanine iodide (DiOC 6 ) and propidium iodide, viable cells show green fluorescence, dead cells have red emission color. The number of viable cells is expressed as a percentage of the total cell number. 100 000 cells in each variant were taken for 100%. Scale bar is 50 μm.

Article Snippet: The zeta-potential of binase-loaded HNTs was monitored using a Zetasizer Nano ZS instrument (Malvern, United Kingdom).

Techniques: Fluorescence, Microscopy, Incubation, Staining, Variant Assay

Journal: Frontiers in Pharmacology

Article Title: Binase Immobilized on Halloysite Nanotubes Exerts Enhanced Cytotoxicity toward Human Colon Adenocarcinoma Cells

doi: 10.3389/fphar.2017.00631

Figure Lengend Snippet: Microscopic visualization of Colo 320 cells after 24 h of incubation with 100 μg/ml HNTs. Dark-field image of cells (left) , fluorescent image (middle) , merge image (right) . Cells nuclei are stained with DAPI.

Article Snippet: The zeta-potential of binase-loaded HNTs was monitored using a Zetasizer Nano ZS instrument (Malvern, United Kingdom).

Techniques: Incubation, Staining

Journal: Nature Communications

Article Title: Structural basis of arrestin-3 activation and signaling

doi: 10.1038/s41467-017-01218-8

Figure Lengend Snippet: IP mediated trimerization and receptor-independent activation in cells. a - b Sedimentation velocity analytical ultracentrifugation (SVAUC) of arrestin-3. Measurements were performed in triplicate. a Representative SVAUC run in the absence of IP predicts a molecular weight matching a monomer. b In the presence of 100 µM IP , the predicted molecular weight is consistent with a trimer. c Representative size exclusion chromatograms of arrestin-3 (Arr3) in the presence and absence of IP . In the absence of IP , 60 µM arrestin-3 (black) elutes at volume corresponding to the Stokes radius of a globular protein with a molecular mass of 64 ± 5.9 kDa. In the presence of 100 μM IP , the elution volume is consistent with a globular protein of molecular mass 169.6 ± 9.7 kDa. The ratio of these molecular weights is consistent with IP -dependent trimer formation. Measurements were performed in triplicate, errors are ± SEM. d Plot of the probability of the distances between spin labels at S13 and A392 for 100 µM arrestin-3 in the presence of the indicated molar ratios of IP ; inset shows the location of the spin labeled sites in basal arrestin-3 (PDB entry 3P2D ). e Comparison of JNK3 activation by GFP-arrestin-3 and Cys-less mutant. GFP-tagged arrestin allowed comparison of the expression levels of wild-type and variant arrestin-3, as described , . JNK3 activation (mean ± SEM) was assessed by measuring the pp-JNK3 levels in COS7 cells co-transfected with HA-ASK1, HA-JNK3 and GFP or Venus-tagged wild-type and Cys-less arrestin-3. Assay was repeated five times and JNK3 phosphorylation was compared by one-way ANOVA followed by Bonferroni post hoc test with correction for multiple comparisons. *** p < 0.001

Article Snippet: The expression level of HA-ASK1, Flag-JNK3 and arrestin-3 were assessed by Western analysis using antibodies against the HA tag (Cell Signaling #C29F4, 1:1000 dilution), the Flag tag (Sigma #F3165, 1:500 dilution), arrestin (F4C1) or GFP (JL-8, Choltech #632381, 1:2000 dilution), respectively.

Techniques: Activation Assay, Sedimentation, Analytical Ultracentrifugation, Molecular Weight, Labeling, Comparison, Mutagenesis, Expressing, Variant Assay, Transfection, Phospho-proteomics

Journal: Journal of Diabetes Research

Article Title: Precedence of Bone Loss Accompanied with Changes in Body Composition and Body Fat Distribution in Patients with Type 2 Diabetes Mellitus

doi: 10.1155/2023/6753403

Figure Lengend Snippet: Demographic data of 596 patients with T2DM.

Article Snippet: The blood samples were collected after fasting for more than 8 h. We measured the levels of fasting plasma glucose, glycosylated hemoglobin A1c (HbA1c) (variant II glycosylated hemoglobin analyzer, Bio-Rad, high-performance liquid chromatography), total cholesterol, triglyceride (TG), high-density lipoprotein cholesterol (HDL-C), and low-density lipoprotein cholesterol (LDL-C) (Siemens ADVIA 2400 automatic biochemical analyzer).

Techniques:

Journal: Journal of Diabetes Research

Article Title: Precedence of Bone Loss Accompanied with Changes in Body Composition and Body Fat Distribution in Patients with Type 2 Diabetes Mellitus

doi: 10.1155/2023/6753403

Figure Lengend Snippet: Linear regression analysis of body composition index, L 1-4 BMD, and FNBMD. Note: adjusted: age, sex, course of T2DM, chronic complications of T2DM, BMI, FBG, HbA1c, TG, LDL-C, HDL-C, SBP, DBP, and medication history. FMI: fat mass index; MMI: muscle mass index; M/F: muscle/fat mass ratio; TFMI: trunk fat mass index; ASMI: appendicular skeletal muscle mass index; A/T: appendicular skeletal muscle mass/trunk fat mass ratio.

Article Snippet: The blood samples were collected after fasting for more than 8 h. We measured the levels of fasting plasma glucose, glycosylated hemoglobin A1c (HbA1c) (variant II glycosylated hemoglobin analyzer, Bio-Rad, high-performance liquid chromatography), total cholesterol, triglyceride (TG), high-density lipoprotein cholesterol (HDL-C), and low-density lipoprotein cholesterol (LDL-C) (Siemens ADVIA 2400 automatic biochemical analyzer).

Techniques:

Journal: Journal of Diabetes Research

Article Title: Precedence of Bone Loss Accompanied with Changes in Body Composition and Body Fat Distribution in Patients with Type 2 Diabetes Mellitus

doi: 10.1155/2023/6753403

Figure Lengend Snippet: Binary logistic regression analysis of body mass index, body composition index, and FNBMD reduction. Note: adjusted: age, sex, course of T2DM, chronic complications of T2DM, BMI, FBG, HbA1c, TG, LDL-C, HDL-C, SBP, DBP, and medication history. FMI: fat mass index; MMI: muscle mass index; M/F: muscle/fat mass ratio; TFMI: trunk fat mass index; ASMI: appendicular skeletal muscle mass index; A/T: appendicular skeletal muscle mass/trunk fat mass ratio.

Article Snippet: The blood samples were collected after fasting for more than 8 h. We measured the levels of fasting plasma glucose, glycosylated hemoglobin A1c (HbA1c) (variant II glycosylated hemoglobin analyzer, Bio-Rad, high-performance liquid chromatography), total cholesterol, triglyceride (TG), high-density lipoprotein cholesterol (HDL-C), and low-density lipoprotein cholesterol (LDL-C) (Siemens ADVIA 2400 automatic biochemical analyzer).

Techniques:

Journal: Brain

Article Title: Genetic landscape of congenital insensitivity to pain and hereditary sensory and autonomic neuropathies

doi: 10.1093/brain/awad328

Figure Lengend Snippet: Novel (likely) pathogenic variants in CIP/HSAN genes

Article Snippet: Datasets were analysed with regard to novel variants in known CIP/HSAN genes [genes from the panel ‘pain syndromes’ v1.12, Genomics England Panel App were prioritized ( https://panelapp.genomicsengland.co.uk/panels/288/ )].

Techniques: Variant Assay

Journal: Brain

Article Title: Genetic landscape of congenital insensitivity to pain and hereditary sensory and autonomic neuropathies

doi: 10.1093/brain/awad328

Figure Lengend Snippet: Novel variants in CIP/HSAN genes classified as likely pathogenic or VUS+ after reclassification

Article Snippet: Datasets were analysed with regard to novel variants in known CIP/HSAN genes [genes from the panel ‘pain syndromes’ v1.12, Genomics England Panel App were prioritized ( https://panelapp.genomicsengland.co.uk/panels/288/ )].

Techniques: Variant Assay