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. 2021 Aug 30;118(36):e2105510118. doi: 10.1073/pnas.2105510118

Fig. 4.

Fig. 4.

Structural mechanism of recognition of the DLF motif from SNX1 by SNX27 FERM. (A) Conservation analysis of the FERM domain of SNX27 using the ConSurf Server (https://consurf.tau.ac.il) shows that the SNX1-binding groove among the F3 submodule is strictly conserved. The surface of SNX27-FERM is colored as light gray (nonconserved) and purple (strictly conserved). SNX1 peptide is shown in cyan sticks. (B) Close-up view showing the detailed binding between the SNX1 DLF motif (cyan) and SNX27-FERM (partially transparent surface; blue and red represent positive and negative charge, respectively). Critical residues on SNX27-FERM important for SNX1 recognition are highlighted in stick mode; black dashed lines indicate the observed hydrogen bonds and salt bridges in the structure. (C) GST, GST-SNX168-90 WT, and mutants (E−5A, Q−3A, D−2A, L−1A, F0A, and DLF) pull down His6-SNX27-FEEM. Shown is a Coomassie Blue–stained SDS/PAGE gel of bound samples of GST in right and GST-SNX1 in left. * indicates the retained His6-SNX27-FERM. (D) Binding affinity between MBP-SNX27-FERM protein and WT or corresponding mutants of SNX168-90 peptides in C determined by ITC. Association constant (Ka) is shown together with errors from data fitting. (E) GST and GST-SNX168-90 pull-down of purified MBP-SNX27-FERM WT, SR, R438A, R438E, QI, and R498A, respectively. Shown is a Coomassie Blue–stained SDS/PAGE gel of bound samples of GST on the right and GST-SNX1 on the left. * indicates the retained MBP-SNX27-FERM. (F) Binding affinity between MBP-SNX27-FERM WT or mutants in E and SNX168-90 WT determined by ITC. Association constant (Ka) is shown together with errors from data fitting.