(A) Sequence alignment of the β1-loop-β2 region in structurally characterized PH domains. Conserved residues are colored: grey shading, glycine; red, positively charged residues; blue, hydrophobic residues. (B) Electrostatic surface of the membrane-bound BBSome–ARL6GTP complex, and close-up views of the BBS5 pleckstrin homology (PH) domains. For the PH1 domain, the canonical (C) and atypical (A) sites for lipid binding are occluded by BBS9 (dark green ribbon) and BBS8 (gold ribbon), respectively. For the PH2 domain, the C site is blocked by BBS9, but the A site is accessible. The lipids at the A and C sites, shown as yellow and cyan sticks, respectively, are diC4-PtdIns(4,5)P2 and are modeled based on the structural alignment of the BBS5 PH domains with the lipid-bound ASAP1 PH domain (PDB ID: 5C79) (C) Structural overlay of the lipids bound to the PH domains listed in the above sequence alignment (PDB IDs: 1MAI, 1W1G, 5C79, 1H10, 2UVM, 2I5C, 1FAO, 1FHW, 1U27), showing the consistency of the lipid position for both PH domains. The lipids are shown as stick models and are overlaid on the PH1 and PH2 domains of BBS5 (orange ribbon).