Fig. 1: PTBP1 and EMCV-IRES-DtoF are flexible in the free state and rigidify within the complex. | Nature Communications

Fig. 1: PTBP1 and EMCV-IRES-DtoF are flexible in the free state and rigidify within the complex.

From: Integrative solution structure of PTBP1-IRES complex reveals strong compaction and ordering with residual conformational flexibility

Fig. 1

a Domain scheme of PTBP1 and an overview of the EMCV-IRES-DtoF RNA construct used in this study. b 2D 1H-15N TROSY spectrum of EMCV-IRES-DtoF imino cross-peak signals confirm base-pairing. Imino cross-peaks of G323 and U334 of SLE shifted as indicated by red arrows. c CW-EPR spectra of SLD and SLF (black) and EMCV-IRES-DtoF (magenta) with spin label attached to nucleotide 309 and 361, respectively. The sharpness of the signals does not change, implying a similar flexibility of the label in the single SL RNA as in the whole EMCV-IRES-DtoF. d The SAXS data measured on free EMCV-IRES-DtoF shows in the Kratky plot (right) the characteristics of a flexible and unstructured molecule. e The SAXS curve of the PTBP1-EMCV-IRES-DtoF complex shows in the Kratky plot (right) characteristics of an overall structured molecule with reduced flexibility compared to the free RNA shown in (d). f Overview of spin-label attachment sites on the individual RRMs of PTBP1 used in this study. Additionally, spin labeling sites were placed in the linker region between RRM1 and RRM2 (152 and 156) and RRM2 and RRM34 (288, 315, 327), respectively. g Selected DEER-EPR measurements using spin labels attached to RRM2 (202, 235), RRM3 (388) and RRM4 (475). In the free state (black), signals decay smoothly (top panel), reflecting a broad distance distribution (bottom panel) with the exception of RRM3-RRM4, which interact stably and, thus, lead to an oscillatory decay corresponding to a narrow distance distribution. Upon complex formation (green), the mean distance between RRM2 and RRM4 can increase (202/475) or decrease (235/475), while there is no significant change upon binding for RRM3/RRM4 (388/475). These distance distributions reflect a more ordered and rigidified arrangement of the RNA within the complex. Semi-transparent areas in distance distribution plots correspond to 95% confidence intervals. The data underlying panels (b–e, g) are provided as Source Data.

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