supply the recognition specificity needed for these interactions, rather than driving the interactions per se. This hypothesis is supported by recent work from Zelzer and coworkers (14) revealing that the swapping of PAS domains between SIM and TRH confers the functional specificity of the PAS domain, rather than that of the parent protein. Based on the experimentally determined structures of PYP light-cycle intermediates (36), we propose that the region of the PAS/PYP fold that is involved in protein-protein interaction is centered around residue 52, as shown in Fig. 5. This hypothesis can be tested by site-directed mutagenesis of residues highlighted in this interface (Fig. 5).

Several PAS-containing proteins bind ligands and/or cofactors. To date, however, mapping the ligand binding to the PAS domain itself has been demonstrated only for the FixL (16, 21, 4) and AHR (15) proteins. The PAS domain of FixL binds heme, whereas the PAS-B domain of AHR binds dioxin and other poly-cyclic aromatic hydrocarbons. Interestingly, for both molecules, the minimum size of the PAS domain that is able to bind the ligand is ~130 residues, the size of the entire PAS/PYP module. Because of a reduction in size of several side chains in ARNT compared to PYP, the 3D model of the ARNT PAS domain displays an internal cavity large enough to accommodate a medium-sized ligand (one-half a heme). Thus, the region previously occupied by the PYP’s chromophore is a logical choice for a ligand pocket.

In summary. PYP appears to exhibit all of the major structural and functional features characteristic of the PAS domain superfamily: a modular domain of ~125–150 residues, a sensor function linked to ligand or cofactor binding, and signal transduction capability governed by heterodimeric assembly. Thus, we propose the testable hypothesis that the entire PYP protein fold is the structural prototype for the modular, 3D, PAS-A and PAS-B domain folds in PAS-containing proteins. This PAS/PYP module provides a structural model to guide experimental testing of hypotheses regarding ligand-binding, dimerization, and signal transduction in PAS proteins.

We thank Christopher Bruns and Christopher D.Putnam for preliminary sequence searches, alignments, and analyses; C.Bruns for help with the illustrations; and C.Bruns, C.D.Putnam, Ulrich K. Genick, and John Tainer for useful criticism and discussion. Research on PYP is funded by the National Institutes of Health Grant GM37684 (to E.D.G.).

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