This review includes an analysis of available X-ray and NMR structures of both members of the immunophilin family; cyclophilins and the FK-506 binding proteins (FKBPs). Available structures are compared and contrasted to highlight different structural features seen both within and between species. Each immunophilin family has been structurally characterised with a variety of small molecule ligands, principally immunosuppressive drugs and their analogues and an overview of these complexes is also presented. Currently the Protein Data Base contains over 60 entries for cyclophilins and over 40 entries for FKBPs. A number of FKBP related structures are also available including structures of MIP (Macrophage Infectivity Potentiator protein) from Legionella pneumophila and Trypanosoma cruzi and Trigger Factor from Mycoplasma genitalium. For all structures discussed in the review a summary of the available biological data is also presented.

Unexpectedly, not interested in sulfur: the antiarthritic complex [Au(PEt3)Cl] (1) reacts with crystals of cyclophilin‐3 (Cyp‐3), a peptidyl prolyl isomerase enzyme linked to cellular stress responses, to form an Nε‐bound [AuPEt3]+ adduct with the active site residue His 133, despite the presence of four Cys thiol groups in the protein. Complex 1 is a potent inhibitor of the enzyme (IC50=14 nM).

Cyclophilin 40 (CyP40) is a tetratricopeptide repeat (TPR)-containing immunophilin and a modulator of steroid receptor function through its binding to heat shock protein 90 (Hsp90). Critical to this binding are the carboxyl-terminal MEEVD motif of Hsp90 and the TPR domain of CyP40. Two different models of the CyP40-MEEVD peptide interaction were used as the basis for a comprehensive mutational analysis of the Hsp90-interacting domain of CyP40. Using a carboxyl-terminal CyP40 construct as template, 24 amino acids from the TPR and flanking acidic and basic domains were individually mutated by site-directed mutagenesis, and the mutants were coexpressed in yeast with a carboxyl-terminal Hsp90␤ construct and qualitatively assessed for binding using a ␤-galactosidase filter assay. For quantitative assessment, mutants were expressed as glutathione S-transferase fusion proteins and assayed for binding to carboxyl-terminal Hsp90␤ using conventional pulldown and enzyme-linked immunosorbent assay microtiter plate assays. Collectively, the models predict that the following TPR residues help define a binding groove for the MEEVD peptide: Lys-227, Asn-231, Phe-234, Ser-274, Asn-278, Lys-308, and Arg-312. Mutational analysis identified five of these residues (Lys-227, Asn-231, Asn-278, Lys-308, and Arg-312) as essential for Hsp90 binding. The other two residues (Phe-234 and Ser-274) and another three TPR domain residues not definitively associated with the binding groove (Leu-284, Lys-285, and Asp-329) are required for efficient Hsp90 binding. These data confirm the critical importance of the MEEVD binding groove in CyP40 for Hsp90 recognition and reveal that additional charged and hydrophobic residues within the CyP40 TPR domain are required for Hsp90 binding.In the absence of hormone, estrogen receptor ␣ is found predominantly in the nucleus as part of a multicomponent complex with Hsp90 1 (1). This abundant molecular chaperone plays an essential role in maintaining steroid receptors in a hormone-activable state, and the association of receptor with Hsp90 is a prerequisite for efficient signal transduction (1). In addition to dimeric Hsp90 and the receptor subunit, mature steroid receptor complexes contain p23 and one of a group of modulating proteins that include the immunophilins cyclophilin 40 (CyP40), FKBP51, and FKBP52 (1). CyP40 was first isolated as a component of estrogen receptor ␣ heterocomplexes (2). This novel cyclophilin binds the immunosuppressant cyclosporin A and shares structural similarity with its partner co-chaperones, FKBP51 and FKBP52, both immunophilins of the FK506-binding class (1). The structure of these large immunophilins is characterized by an amino-terminal immunophilin-like domain that is targeted by immunosuppressant drugs and overlaps a peptidylprolyl isomerase function and a carboxyl-terminal protein interaction domain incorporating 3 units of the tetratricopeptide repeat (TPR) motif (3). These immunophilins have been implicated in steroid receptor function through their association with Hsp...

The X-ray structures are consistent with the role of Cyclophilin 40 as a multifunctional signaling protein involved in a variety of protein-protein interactions. The intermolecular helix-helix interactions in the tetragonal form mimic the intramolecular interactions found in the fully folded monoclinic form. These conserved intra- and intermolecular TPR-TPR interactions are illustrative of a high-fidelity recognition mechanism. The two structures also open up the possibility that partially folded forms of TPR may be important in domain swapping and protein recognition.

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During cut-and-paste mariner/Tc1 transposition, transposon DNA is cut precisely at its junction with flanking DNA, ensuring the transposon is neither shortened nor lengthened with each transposition event. Each transposon end is flanked by a TpA dinucleotide: the signature target site duplication of mariner/Tc1 transposition. To establish the role of this sequence in accurate DNA cleavage, we have determined the crystal structure of a pre-second strand cleavage mariner Mos1 transpososome. The structure reveals the route of an intact DNA strand through the transposase active site before second strand cleavage. The crossed architecture of this pre-second strand cleavage paired-end complex supports our proposal that second strand cleavage occurs in trans. The conserved mariner transposase WVPHEL and YSPDL motifs position the strand for accurate DNA cleavage. Base-specific recognition of the flanking DNA by conserved amino acids is revealed, defining a new role for the WVPHEL motif in mariner transposition and providing a molecular explanation for in vitro mutagenesis data. Comparison of the pre-TS cleavage and post-cleavage Mos1 transpososomes with structures of Prototype Foamy Virus intasomes suggests a binding mode for target DNA prior to Mos1 transposon integration.