Journal: iScience

Article Title: Charge Characteristics of Agouti-Related Protein Implicate Potent Involvement of Heparan Sulfate Proteoglycans in Metabolic Function

doi: 10.1016/j.isci.2019.10.061

Figure Lengend Snippet: Glycan Array Analysis of AgRP Binding to Heparan Sulfate Oligosaccharides (A) Fluorescent image of the glass slide glycan arrays showing fluorescence signals (green dots) of AgRP binding to 52 immobilized heparan sulfate oligosaccharides (low-molecular-weight heparan sulfate, LMHS). (B) Bar graph showing the relative fluorescence intensity of AgRP binding to the heparan sulfate oligosaccharides arrays. Heparan sulfate oligosaccharides 22 and 41 show the highest intensity. (C) The structures of the different heparan sulfate oligosaccharides on the slide microarray in A. Data are represented as mean ± SEM.

Article Snippet: Amine(-NH 2 )-linked heparan sulfate glycan compounds (Glycan Therapeutics) were immobilized on NHS-activated surface-coated slides (Nexterion Slide-H, Applied Microarrays) using a robotic microarray printer (Microgrid II, Digilab) that was equipped with StealthSMP4B microarray pins (Telechem) to couple heparan sulfate glycan compounds by covalent binding via (-NH 2 ) reactive chemistry.

Techniques: Glycoproteomics, Binding Assay, Fluorescence, Molecular Weight, Microarray

Journal: Nucleic Acids Research

Article Title: The Effects of Sequence Variation on Genome-wide NRF2 Binding—New Target Genes and Regulatory SNPs

doi: 10.1093/nar/gkw052

Figure Lengend Snippet: Testing the limits of NRF2 binding. Systematic multivariate ARE results by protein binding microarray for NRF2-MAFG heterodimer are shown. Tolerance for sequence variation was studied for positions 1–3 (white), 6–8 (grey) and 9–11 (dark grey) ( A ) and positions 1, 5 and 11 (seed sequences are shown in grey) ( B ). Results are depicted as measured binding relative to NQO1.ARE binding (black) (mean ± S.E.M., n = 45).

Article Snippet: The dilutions were dispensed in a 384-well plate (polypropylene plate No 267462, Nunc, N.Y, USA) and printed onto the avidin-coated glass slides with a microarray printer (BioRobotics MicroGrid II, BioRobotics Ltd, Cambridge, UK).

Techniques: Binding Assay, Protein Binding, Microarray, Sequencing

Journal: Nucleic Acids Research

Article Title: The Effects of Sequence Variation on Genome-wide NRF2 Binding—New Target Genes and Regulatory SNPs

doi: 10.1093/nar/gkw052

Figure Lengend Snippet: A SNP in FTL promoter has drastic effects of NRF2 binding and transcriptional activation. ( A ) A promoter analysis of the FTL gene at chr19 showing the location of experimentally verified NRF2 binding ARE together with dbSNP (v138) and ENCODE ChIP-seq data. ChIP-seq track displays combined MAFF and MAFK binding signals in H1-hESC (MAFK), K562 (MAFF, MAFK), HeLa-S3 (MAFK), HepG (MAFF, MAFK) and IMR90 (MAFK) cell lines. ( B ) Detailed view showing FTL ARE sequence and the SNP (rs113067944, A→C) position. ( C ) Protein binding microarray results for FTL.ARE.A and the SNP bearing FTL.ARE.C. Results are calculated as measured binding relative to NQO1.ARE binding (mean ± S.E.M, n = 39.). Scramble oligonucleotides served as negative control. ( D ) HEK-293T cells were transfected with NQO1-ARE and FTL-ARE bearing either allele A or allele C with and without NRF2 -expressing plasmids. Twenty-four h after transfection cells were treated with NRF2 inducer (L-SFN) for 16 h followed by luciferase activity measurements. An empty pGL3 promoter vector served as control and activities were normalized to β-galactosidase activity. Results are shown relative to control (mean± S.E.M, n = 4).

Article Snippet: The dilutions were dispensed in a 384-well plate (polypropylene plate No 267462, Nunc, N.Y, USA) and printed onto the avidin-coated glass slides with a microarray printer (BioRobotics MicroGrid II, BioRobotics Ltd, Cambridge, UK).

Techniques: Binding Assay, Activation Assay, ChIP-sequencing, Sequencing, Protein Binding, Microarray, Negative Control, Transfection, Expressing, Luciferase, Activity Assay, Plasmid Preparation, Control