Figure 4.
Effect of R70, R111, N104, Y103 to the binding of BC1 RNA. (A–C) Docking models of the FMRP-NT WT, FMRP-NT R70E-R111E and FMRP-NT Y103A-N104A interaction with BC1, respectively. The yellow RNA backbone represents the nucleotides involved in the interaction. The key amino acids detected by mass spectrometry analysis are depicted as blue sticks. (D) EMSA experiment using BC1 RNA, FMRP-NT WT and FMRP-NT R70E-R111E mutant (10–100 ng). Shifted and unbound RNA are indicated by asterisk and arrowhead, respectively; the histogram shows the ratio of the Kd of mutant FMRP-NT versus the Kd of WT FMRP-NT. Error bars represent SE: **P < 0.01, Student's test, n = 3. (E) Competition experiments using unlabelled BC1 (lanes 3–5) and U1 (lanes 6–8) transcripts. (F) and (G) The same as in (D) and (E) but using the Y103A-N104A mutant. P = 0.07 Student's test, n = 5.