Journal: STAR Protocols

Article Title: Adverse outcome pathway-based analysis of liver steatosis in vitro using human liver cell lines

doi: 10.1016/j.xpro.2023.102500

Figure Lengend Snippet:

Article Snippet: Tris-HCl (Cas-no.: 1185-53-1) Caution: Tris-HCl is classified as irritant , PanReac AppliChem , Cat. no. A1087.

Techniques: Recombinant, Modification, Spectroscopy, Transfection, Software, Cell Culture, Suction Filtration, Cell Counting, Inverted Microscopy, Transferring, Mass Measurement, Spectrophotometry, Real-time Polymerase Chain Reaction

Journal: Scientific Reports

Article Title: Structure and nucleic acid binding properties of KOW domains 4 and 6–7 of human transcription elongation factor DSIF

doi: 10.1038/s41598-018-30042-3

Figure Lengend Snippet: Solution structure statistics for KOW4 (S522-G647) and KOW6-7 (G961-A1087).

Article Snippet: Although the twin KOW6-7 domains are positioned near the exiting RNA in the cryo-EM structure of the RNAP II/DSIF complex , we could not detect nucleic acid binding for the corresponding KOW6-7 (G961-A1087) construct, indicating that it probably interacts exclusively with protein factors necessary for RNA elongation and/or RNA processing during transcription termination (Fig. ).

Techniques: Derivative Assay

Journal: Scientific Reports

Article Title: Structure and nucleic acid binding properties of KOW domains 4 and 6–7 of human transcription elongation factor DSIF

doi: 10.1038/s41598-018-30042-3

Figure Lengend Snippet: Solution structure of KOW6-7 (G961-A1087). ( a ) Superposition of the 20 lowest energy structures from Q974-A1087. The region from G961 to E973 is unstructured and not shown. ( b ) Ribbon representation highlighting KOW6 β1–β5) in cyan, and KOW7 (β6–β10) in red ( c ) Stick representation showing relevant amino acids of the domain interface. ( d ) Distribution of R 1 / R 2 for KOW6-7 (G961-A1087). Colour assignment as in (b).

Article Snippet: Although the twin KOW6-7 domains are positioned near the exiting RNA in the cryo-EM structure of the RNAP II/DSIF complex , we could not detect nucleic acid binding for the corresponding KOW6-7 (G961-A1087) construct, indicating that it probably interacts exclusively with protein factors necessary for RNA elongation and/or RNA processing during transcription termination (Fig. ).

Techniques:

Journal: Scientific Reports

Article Title: Structure and nucleic acid binding properties of KOW domains 4 and 6–7 of human transcription elongation factor DSIF

doi: 10.1038/s41598-018-30042-3

Figure Lengend Snippet: Rotational diffusion tensor analysis for KOW6-7 (G981-A1087) (79 vectors).

Article Snippet: Although the twin KOW6-7 domains are positioned near the exiting RNA in the cryo-EM structure of the RNAP II/DSIF complex , we could not detect nucleic acid binding for the corresponding KOW6-7 (G961-A1087) construct, indicating that it probably interacts exclusively with protein factors necessary for RNA elongation and/or RNA processing during transcription termination (Fig. ).

Techniques: Diffusion-based Assay

Journal: Scientific Reports

Article Title: Structure and nucleic acid binding properties of KOW domains 4 and 6–7 of human transcription elongation factor DSIF

doi: 10.1038/s41598-018-30042-3

Figure Lengend Snippet: Structural overlay of hSpt5 KOW domains and sequence comparison with the consensus KOW motif. ( a ) The structures of hSpt5 KOW2 (PDB: 2E6Z), KOW3 (PDB: 2DO3), KOW4 (S522-G647) (this work, PDB: 6EQY), KOW5 (PDB: 2E70), and KOW6-7 (G961-A1087) (this work, PDB: 6ER0) were used for the overlay. ( b ) Comparison of the hSpt5 KOW sequences with the consensus KOW motif highlights the conserved residues in yellow and orange and the insertion in KOW6 in bold.

Article Snippet: Although the twin KOW6-7 domains are positioned near the exiting RNA in the cryo-EM structure of the RNAP II/DSIF complex , we could not detect nucleic acid binding for the corresponding KOW6-7 (G961-A1087) construct, indicating that it probably interacts exclusively with protein factors necessary for RNA elongation and/or RNA processing during transcription termination (Fig. ).

Techniques: Sequencing

Journal: Scientific Reports

Article Title: Structure and nucleic acid binding properties of KOW domains 4 and 6–7 of human transcription elongation factor DSIF

doi: 10.1038/s41598-018-30042-3

Figure Lengend Snippet: Schematic representation of Spt4 and Spt5 proteins. Bacterial NusG consists of an N-terminal domain (NTD) followed by one KOW domain. The archaeal dimeric Spt4/5 consists of a NusG N-terminal (NGN) domain in Spt5, which is homologous to NusG and interacts with Spt4, and one KOW domain. Eukaryotic Spt4/5 and the human DSIF (hSpt4/5) harbour an additional N-terminal acidic region that is followed by the NGN domain. KOW1-5 are found in all eukaryotes, however the CTRs and the C-terminal region adjacent to KOW5 are only present in the Spt5 of metazoans and plants. Numbers represent amino acid positions, dotted lines indicate the regions of the KOW4 and KOW6-7 variants used in this study.

Article Snippet: Although the twin KOW6-7 domains are positioned near the exiting RNA in the cryo-EM structure of the RNAP II/DSIF complex , we could not detect nucleic acid binding for the corresponding KOW6-7 (G961-A1087) construct, indicating that it probably interacts exclusively with protein factors necessary for RNA elongation and/or RNA processing during transcription termination (Fig. ).

Techniques:

Journal: Scientific Reports

Article Title: Structure and nucleic acid binding properties of KOW domains 4 and 6–7 of human transcription elongation factor DSIF

doi: 10.1038/s41598-018-30042-3

Figure Lengend Snippet: Determination of nucleic acid binding affinities by fluorescence anisotropy measurements. 50 nM ( a , d ) or 25 nM ( b , c ) of fluorescent labelled nucleic acids as indicated were titrated with ( a ) KOW4 (S522-G647), ( b ) KOW4 (G531-L705), ( c ) KOW4 (G531-G754), ( d ) KOW6-7 (G961-A1087). The curves in ( a ) represent the best fit to a two-component binding equation to determine the K D values for ssDNA1 (12.2 ± 0.6 µM), dsDNA1 (20.1 ± 0.4 µM), ssDNA2 (24.5 ± 3.2 µM), dsDNA2 (41.2 ± 9.0 μM), and ssRNA (39.5 ± 7.4 μM).

Article Snippet: Although the twin KOW6-7 domains are positioned near the exiting RNA in the cryo-EM structure of the RNAP II/DSIF complex , we could not detect nucleic acid binding for the corresponding KOW6-7 (G961-A1087) construct, indicating that it probably interacts exclusively with protein factors necessary for RNA elongation and/or RNA processing during transcription termination (Fig. ).

Techniques: Binding Assay, Fluorescence