Crystallogr. one incomplete module (LRR12). The consensus residues in the LRR invariant fragment are shown above the sequence in standard font; the consensus residues in the variable fragment are shown above the sequence in italics. Text color corresponds to that shown in panel B. Underlined residues are the basic residues that define a positively charged cluster at the ascending loops of SDS22 (1R,2S)-VU0155041 (see F and G). (B). SDS22 structure shown as cartoon and colored to illustrate the individual LRR caps and LRR modules, as indicated in panel A. LRR repeats are numbered. The consensus residues in the LRR invariant fragment (LxxLxLxxNxI) are shown as sticks; the conserved leucines define the hydrophobic core while the conserved asparagines mediate inter-repeat hydrogen bonds. (C) LRR5 shown as a cartoon with the consensus residues shown as sticks. (D) SDS22 colored as in panel B and rotated to illustrate the curvature of the protein. (E) The N- (right) and C-terminal (left) caps of SDS22, with residues mediating hydrogen bonds shown as sticks and labeled. (F) Electrostatic surface of SDS22 shown in the same orientation as in panel B (top) and rotated by 90 (bottom). (G) The residues that define the strong basic patch in SDS22. Five sulphate ions, which were present in the crystallization mother liquor, are shown as sticks. The LRR superstructure is stabilized by an extensive hydrophobic core comprised of conserved, closely packed (iso)leucines; it is also stabilized by inter-repeat backbone-backbone and backbone-sidechain (i.e., the asparagine ladder) hydrogen bonds (Figure 1B). Finally, the first and last LRR repeats are shielded from solvent by an N- and a C-terminal cap, respectively (Figure 1E). The N-terminal cap is an -helix defined (1R,2S)-VU0155041 by residues 69-75 while the C-terminal cap (1R,2S)-VU0155041 is a canonical LRR-cap motif comprised of residues 339-354. The LRR-cap forms an amphipathic -helix that aligns with the 310-helix of LRR11, and is followed by a -turn and an extended conformation that forms hydrogen bonds with the -strand of LRR12 and the last six residues of SDS22. The LRR-cap is further stabilized by side chain hydrogen bonds between Tyr339SDS22 and Asp354SDS22, and between Arg340SDS22 and Thr356SDS22. A distinctive feature of SDS22 is the presence of a large negatively charged patch on its concave side (Figure 1F). A second negatively FRPHE charged patch is formed by the convex side and is caused by repeating glutamate residues at position 15 of LRRs 1C6 and 8C10. However, its most distinctive feature is an extended, highly basic patch that is generated by seven arginine/lysine residues (SDS22 residues Lys133, Arg155, Lys175, Arg197, Arg199, Lys219 and Arg241) that belong to the ascending (1R,2S)-VU0155041 loops of LRRs 3C8. In the crystal, these residues coordinate five sulfate ions (Figures ?(Figures1G,1G, S2B and S2C). Model of the PP1:SDS22 complex Using the docking program HADDOCK (Dominguez et al., 2003; De Vries et al., 2007), we generated a computational model of the PP1:SDS22 complex (Figures 2A and 2B). Based on this model, the positively charged PP1 helices 5/6 and the 7-8 loop of PP1 are predicted to interact with the acidic concave surface of SDS22, creating a large buried surface area of 2500 ?2. Twelve residues of SDS22, distributed over nine LRRs, are predicted to bind directly to PP1. Most of these residues are part of the extended -sheet of SDS22. SDS22 binding clearly masks a positively charged surface area of PP1 but does not create an obvious additional surface groove towards the catalytic site of PP1 (Figure S2D)..