Figure 5. Structural and Mutational Analyses of the KlEst1-KlCdc13_EBM Interaction.

(A) Domain organization of KlEst1 and KlC\Cdcl3. Domains of KlEst1 (TPR, tetratricopeptide repeat; HHD, helical hairpin domain; IM, insertion motif) are colored in green, cyan, and slate blue, respectively, and KlCdcl3_EBM is in yellow. The shaded area indicates the interaction between KlEst1 and KlCdcl3_EBM- (DBD: DNA-binding domain) (B) ITC measurements of interactions between KlEstl with different fragments of KlCdcl3 within KlCdcl3_RD. Inset: ITC titration data. UD: undetectable. (C) Overall structure of the KlEstl-KlCdc13 complex. KlEstl and KlCdcl3_EBM are colored as in (A). KlCdcl3_EBM is shown as a stick model. (D) Multiple sequence alignment of Cdcl3_EBM from different yeast species. The conserved residues are highlighted in red and KlCdcl3_EBM-N and KlCdcl3_EBM-c are highlighted in blue boxes. Glu233 in KlCdc13 (equivalent to Glu252 in ScCdcl3) is denoted with an arrow. (E) Electrostatic surface potential of the KlCdc13_EBM-binding cavity of KlEst1 (positive potential, blue; negative potential, red). KlCdc13_EBM is shown in a stick model and colored in yellow. (F) Details of interactions between KlCdc13_EBM-N and its surrounding residues of KlEst1. Hydrogen bonds are denoted as dashed magenta lines. (G) ITC data of the wild-type and mutant KlEst1-KlCdc13_EBM interactions. Counterpart residues in S. cerevisiae proteins are shown in parenthesis. (H) Details of interactions between KlCdc13_EBM-C and its surrounding residues of KlEst1. Glu233 in KlCdc13 and Lys467 in KlEst1 are highlighted with boxes.

See also Figure S5, S6 and Table S3.