Journal: bioRxiv
Article Title: SpoVG and the Kre-ComK Regulatory Module Orchestrate Production of the EPE Toxin in Bacillus subtilis
doi: 10.64898/2026.04.02.716078
Figure Lengend Snippet: SPR analysis of Kre binding to full-length and truncated epeX RNA transcripts. a) Schematic overview of truncated epeX RNA fragments. I ) The full-length RNA construct ( epeX 1–11) spans from the +1 transcriptional start site of epeX to the start codon of epeE (232 nt). II ) To identify specific binding regions, three mid-sized fragments (∼80 nt each) were generated: epeX 1–3, epeX 5–7, and epeX 9–11. III ) For higher-resolution mapping, a tiling strategy was applied using eleven 40 bp fragments ( epeX 1 through epe X 11), each overlapping its neighbouring fragment by 20 bp. b-e) Sensorgrams (left panels) depict response units of Kre to increasing concentrations of epeX RNA fragments (black: 10 nM; red: 25 nM; light green: 50 nM; yellow: 100 nM; blue: 250 nM; turquoise: 500 nM; light blue: 1000 nM; grey: 2500 nM; brown: 5000 nM). Interaction Maps (central panels) show the distribution of association and dissociation rate constants as stability (dissociation rate; log k d ) and recognition (association rate, log k a ). The corresponding calculated sensorgrams from the IM peaks (right panels) are presented on the right, the colours of the curved correspond to the respective peaks from the IM analyses. The data were quantified and the respective overall affinities (K D ) calculated from the respective association ( k a ) and dissociation rates ( k d ) are shown, as well as the peak weights (PW) showing the overall contribution of the respective interaction towards the total sensorgrams given in (%). (see for details). Since bulk binding are not included in the calculations, 100% were not reached in total. The full-length epeX RNA b) epeX 1-11, as well as truncated versions of the epeX RNA c) epeX 1-3, d) epeX 5-7, and e) epeX 9-11, were applied for SPR as well as IM analysis.
Article Snippet: For this purpose, the SPR sensorgrams were exported from the Biacore T200 Evaluation Software 3.2.1 as *.txt files and imported into TraceDrawer Software 1.10.1 (Ridgeview Instruments, Uppsala, Sweden).
Techniques: Binding Assay, Construct, Generated
