Journal: mLife

Article Title: The GntR/VanR transcription regulator AlkR represses AlkB2 monooxygenase expression and regulates n ‐alkane degradation in Pseudomonas aeruginosa SJTD‐1

doi: 10.1002/mlf2.70004

Figure Lengend Snippet: The AlkR protein can efficiently bind to the promoter region of the alkB 2 gene in vitro and the binding can be released by two long‐chain fatty acyl‐CoA compounds. (A) Schematic diagram of the location and the intergenic region of alkB2 and alkR genes. The ‒10 and ‒35 regions of the alkB2 promoter and the transcriptional start site (+1) of the alkB2 gene are highlighted. The three DNA fragments of different lengths in the alkB 2 promoter (P1‐P3) are shown, ranging from positions –50 bp, –76 bp, and –187 bp to the start codon ATG of the alkB 2 gene. (B) Recombinant AlkR could bind to the three DNA fragments (P1, P2, P3) in a concentration‐dependent manner. The EMSA assays were performed at various AlkR/DNA molar ratios, with AlkR/P1 (0.2, 1.0, 2.0, 10.0, 20.0, from left to right), AlkR/P2 (0.1, 0.2, 1.0, 5.0, 10.0, from left to right), and AlkR/P3 (0.2, 1.0, 2.0, 5.0, 10.0, 20.0, from left to right). The BSA protein was used as a negative control (–). The gray triangles represent the shift bands of unbound DNA fragments, and the black triangles represent different shift bands of the protein‐bound DNA fragments. (C) Two long‐chain fatty acyl‐CoA compounds (C16‐CoA and C18‐CoA) could release the binding of AlkR to the P3 fragment. Two long‐chain fatty acyl‐CoA (C16‐CoA or C18‐CoA, 5.0, 10.0, 20.0, 40.0, 80.0, and 150.0 μM, from left to right) were added to the binding system with AlkR (5.8 × 10 −7 M) and P3 fragments (5 × 10 −7 M), and then assessed by EMSA. The reactions supplied without an effector or supplied with acetyl‐CoA (600 μM) were used as a negative control (–) and a positive control (+), respectively. The profiles of 12% PAGE from three independent assays are shown. The gray triangles represent the shift bands of unbound DNA fragments and the black triangles represent different shift bands of the protein‐bound DNA fragments.

Article Snippet: The fragment of the promoter region of the alkB 2 gene was amplified using the primers tagged with biotin via an N33‐TEG‐COOH linker (MedChemExpress LLC) and immobilized with streptavidin‐coated magnetic particles.

Techniques: In Vitro, Binding Assay, Recombinant, Concentration Assay, Negative Control, Positive Control

Journal: eLife

Article Title: Bacterial exonuclease III expands its enzymatic activities on single-stranded DNA

doi: 10.7554/eLife.95648

Figure Lengend Snippet:

Article Snippet: Peptide, recombinant protein , SSB ( E. coli ) , Beyotime Biotech. Inc, China , P7415 , .

Techniques: Recombinant, Plasmid Preparation, Expressing, Chromatography, Protein Purification, Cloning, In Vitro, Amplification, Fluorescence, Sequencing, Software