Figure 5
From: Discovery of Small Molecules that Inhibit the Disordered Protein, p27Kip1
2D 1H-15N TROSY-HSQC NMR analysis of displacement of 15N-p27-D2 from Cdk2/cyclin A by SJ403.
2D 1H-15N TROSY-HSQC NMR spectra of p27-D2/Cdk2/cyclin A in the absence (control) and presence of SJ403 (3 mM), respectively, were recorded. For the residues reported, both free and bound p27-D2 resonances were observed in the TROSY-HSQC spectra. An increase in the relative intensity of free p27-D2 resonances was observed in the TROSY-HSQC spectrum of p27-D2/Cdk2/cyclin A in the presence of SJ403, with respect to the control spectrum (a), while the resonances of p27-D2 bound to Cdk2/cyclin A showed a decrease in relative intensity (b). This permits the calculation of populations of free (f) and bound (b) p27-D2, in the absence (w/o SJ403) and presence of SJ403 (w/SJ403)). The pfw/o SJ403 values represent the relative populations of resonances for free p27-D2 before addition of SJ403 and the pfw/SJ403 values are the relative populations of free state p27-D2 resonances after SJ403 addition. Correspondingly, the pbw/o SJ403 values represent the relative populations of resonances for p27-D2 bound to Cdk2/cyclin A before addition of SJ403 and the pbw/SJ403 values are the relative populations of bound state p27-D2 resonances after SJ403 addition. The mean values and standard deviations of the means for triplicate measurements are shown; one set of triplicate 2D 1H-15N TROSY-HSQC spectra are illustrated in Suppl. Fig. 7.
