Figure 1.
Structures of KLC1-TPR and KLC2-TPR.
(A) A ribbon diagram of KLC1-TPR with inner helix-A (blue), outer helix-B (red), loops (green), and the non-TPR insert (gray). (B) A ribbon diagram of KLC2-TPR. (C) Structural alignment of KLC1-TPR (orange) and KLC2-TPR (green). (D) A ribbon diagram of the highly mobile non-TPR inserts of KLC1-TPR (olive) and KLC2-TPR (dark green). (E) A structure-based sequence alignment of KLC1-TPR and KLC2-TPR using ESPript [38].
Table 1.
Data collection and refinement statistics of KLC1-TPR and KLC2-TPR.
Table 2.
Secondary structure alignments with other known TPR domains.
Figure 2.
Structural alignment of KLC1-TPR with p67phox and HOP1.
(A) KLC1-TPR (orange) is aligned with the p67phox TPR domain (marine) in complex with Rac1 (Yellow). (B) KLC1-TPR (orange) is aligned with the HOP1 TPR domain (marine) in complex with Hsc70 peptide (Yellow). Residues lining the interaction interface of the p67phox and HOP1 TPR domains and their respective binding partners as well as the corresponding KLC1-TPR residues are shown in the circle.
Figure 3.
Binding experiments of the KLC1 and KLC2 TPR domains with the JIP1 peptide.
Isothermal titration calorimetry measurements of (A) KLC1-TPR with the JIP1 peptide, (B) KLC2-TPR with the JIP1 peptide, (C) KLC1-N343S mutant with the JIP1 peptide.
Figure 4.
Comparison of KLC1-TPR and KLC2-TPR polar patches.
(A) N343 of KLC1-TPR interacts with neighboring helix residues T383 and N386 as well as the N343 carboxamide group is available at the surface. (B) S328 of KLC2 is unable to interact with neighboring helix residues due to its short side chain.
Table 3.
Thermodynamic parameters of the ITC experiment between KLC proteins with JIP1 and ALC1 peptides.
Figure 5.
Binding experiments of the KLC1-TPR mutants with ALC1 peptide and protein.
Isothermal titration calorimetry measurements of (A) KLC1-N343S mutant with the ALC1 peptide, (B) KLC1-N301A mutant with the ALC1 peptide, (C) KLC1-TPR with the cytosolic domain of ALC1 protein.
Figure 6.
Comparison of ALC1- and JIP1- binding polar patches of KLC1-TPR.
The distance between N301–N344 clamps of the ALC1-binding polar patch (marine) is 5.7 Å, while the N343-N386 clamp of the JIP1-binding polar patch (yellow) is closer at 3.6 Å. A lysine (orange) is present in the JIP1-binding polar patch compared to an alanine (cyan) of ALC1-binding polar patch.