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. 2020 Jul 30;11:3807. doi: 10.1038/s41467-020-17647-x

Fig. 5. Charged regions in the RNF4 linker promote a compact but not collapsed shape.

Fig. 5

a hydropathy plot of RNF4 revealing the charge characteristics of the amino-acid side chains. The SIMs and RING domain are highlighted, along with the charged linker that connects these two domains. b layout of the RNF4N peptide used for single-molecule analysis. Variants of the RNF4N peptide were produced removing either the basic or acid charge from the SIMs/RING-domain linker region. The mutations used are displayed against the WT peptide. c size exclusion chromatography profile of the WT RNF4N peptide (black) along with ∆Basic (blue) and ∆Acidic (magenta) variants. d single-molecule FRET histograms obtained from RNF4N peptides labelled with donor/acceptor FRET dyes at C-terminus and basic region (labelled residue numbers inset). Arrows inset highlight the FRET efficiency average for WT (black), ∆Basic (blue) and ∆Acidic (magenta) RNF4N peptides. Single-molecule histograms were built from more than 400 molecules. e Distance measurements over time for five independent simulations between acidic residues in the SIM2/3 linker (defined by E59) and the midpoint of the arginine-rich motif for RNF4N WT (top) and ∆Basic (bottom). f Conformational clustering of RNF4N peptides (WT top; ∆Basic bottom) generated from residue-residue contacts over the final 100 ns of each run trajectory from e. The SIMs (orange), arginine-rich region (blue) and acidic region in the SIMs/RING-domain linker (red) are highlighted. Source data are provided as a Source Data file.