Crystal structures of cyclophilin A complexed with cyclosporin A and N-methyl-4-[(E)-2-butenyl]-4,4-dimethylthreonine cyclosporin A

Ke, Hengming; Mayrose, Dale; Belshaw, Peter J.; Alberg, David G.; Schreiber, Stuart L.; Chang, Zhi Yuh; Etzkorn, Felicia A.; Ho, Susanna I.; Walsh, Christopher T.

doi:10.1016/s0969-2126(00)00006-x

Cited by 83 publications

(61 citation statements)

References 50 publications

(93 reference statements)

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“…The conformations of CyPA, CsA, CNA, and CNB in the CyPACsA-CN complex are similar to the corresponding subunits in the CyPA-CsA binary complex (19,23), unbound CN (17), and the FKBP-FK506-CN complex (16,17). Superposition of the C ␣ atoms of the corresponding subunits revealed average displacements of 0.39 Å for CyPA between the binary CyPA-CsA and tertiary CyPA-CsA-CN complexes, and 0.66 and 0.79 Å for CNA and CNB between CyPA-CsA-CN and unligated CN, indicating no substantial change of overall molecular conformations upon formation of the CyPA-CsA-CN complex.…”

Section: Resultsmentioning

confidence: 76%

“…The crystal in the space group P3 2 21 contains one complex of CyPA-CsA-CN in the crystallographic asymmetric unit. The CyPA-CsA-CN structure in P3 2 21 was solved by molecular replacement program AMORE using individual subunits of CyPA (19) and CNA and CNB from the structure of FKBP-FK506-CN (16,17). The orientation of CNA was first located, yielding a correlation coefficient of 0.37 and R factor of 0.39 for the data between 4-and 8-Å resolution.…”

Section: Methodsmentioning

confidence: 99%

“…5). In addition, four pairs of hydrogen bonds between CsA and CyPA: MeBmt-1⅐⅐⅐Gln-63, Abu-2⅐⅐⅐Asn-102, MeLeu-9⅐⅐⅐Trp-121, and MeLeu-10⅐⅐⅐Arg-55, are conserved in both binary CyPA-CsA (19) and ternary CyPA-CsA-CN complexes. However, the side chains of CN-binding moiety of CsA undergo significant conformational changes.…”

Section: Subtle Conformational Changes Of Csa Upon Complex Formation Andmentioning

confidence: 99%

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Crystal structure of calcineurin–cyclophilin–cyclosporin shows common but distinct recognition of immunophilin–drug complexes

Huai

Kim

Liu

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

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253

191

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Calcineurin, a Ca 2؉ ͞calmodulin-dependent protein phosphatase, is the common target for two immunophilin-immunosuppressant complexes, cyclophilin A-cyclosporin A (CyPA-CsA) and FKBP-FK506. How the two structurally distinct immunophilin-drug complexes bind the same target has remained unknown. We report the crystal structure of calcineurin (CN) in complex with CyPA-CsA at 2.8-Å resolution. The CyPA-CsA complex binds to a composite surface formed by the catalytic and regulatory subunits of CN, where the complex of FK506 and its binding protein FKBP also binds. While the majority of the CN residues involved in the binding are common for both immunophilin-immunosuppressant complexes, a significant number of the residues are distinct. Unlike immunosuppressants ͉ FK506 ͉ FK506-binding protein ͉ protein phosphatase ͉ T cell C alcineurin (CN) is a unique serine͞threonine protein phosphatase (PP3 or PP2B) in its regulation by the second messenger Ca 2ϩ and calmodulin. CN is involved in many biological processes, including immune responses (1-5), the second messenger cAMP pathway (3, 6), Na ϩ ͞K ϩ ion transportation in the nephron (7), cell cycle progression in lower eukaryotes (8), cardiac hypertrophy (9), and memory formation (10, 11). The most extensively studied function of CN is its role in the signal transduction pathway during T cell activation. CN has been shown to be a common receptor for two immunophilinimmunosuppressant complexes, cyclophilin A-cyclosporin A (CyPA-CsA) and FK506-binding protein (FKBP)-FK506 (12, 13). The binding of CyPA-CsA or FKBP-FK506 inhibits the calcium-dependent dephosphorylation of the transcription factor nuclear factor of activated T cell (NFAT) by CN, thus blocking T cell receptor-mediated cytokine transcription and T-cell activation (14,15).Since the discovery of CN as a common target for both CyPA-CsA and FKBP-FK506 complexes, how two structurally distinct immunophilin-immunosuppressant complexes recognize a common target has remained a mystery. While the crystal structures of CN and its complex with FKBP-FK506 have shed light on the molecular recognition of CN by 17), the CyPA-CsA-CN structure has remained elusive. We report here the crystal structure of CyPA-CsA-CN at 2.8-Å resolution. The structure revealed that CyPA-CsA and FKBP-FK506 are bound to the same surface of CN, thus providing a molecular basis for understanding the regulation of CN activity by the immunophilin-immunosuppressant. MethodsProtein Purification. The catalytic subunit (CNA, 61 kDa) and regulatory subunit (CNB, 19 kDa) of the ␣ isoform of human CN were coexpressed in Escherichia coli and purified by using a previously described procedure with slight modifications (18). The recombinant human CyPA was expressed as a glutathione S-transferase (GST) fusion protein in E. coli BL21 and purified by glutathione-agarose beads (Sigma). The CyPACsA-CN complex was prepared by a four-step procedure: (i) purification of recombinant human CN by Ni NTA (Qiagen, Valencia, CA) and Q-Sepharose (Pharmacia) columns, (ii) purific...

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Section: Resultsmentioning

confidence: 76%

Section: Methodsmentioning

confidence: 99%

Section: Subtle Conformational Changes Of Csa Upon Complex Formation Andmentioning

confidence: 99%

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Crystal structure of calcineurin–cyclophilin–cyclosporin shows common but distinct recognition of immunophilin–drug complexes

Huai

Kim

Liu

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

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253

191

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“…The overall architecture of CsCyp is highly similar to that of the C. elegans Cyp3 and human CypA structures in complex with CsA (Pflügl et al, 1993;Ke et al, 1994;Dornan et al, 1999) and comprises an eight-stranded antiparallel b-barrel capped at either end by two a-helices (Fig. 2, A and B).…”

Section: On the Structural (Dis)similarities Between Classical And DImentioning

confidence: 81%

“…The CsCyp active site, composed of 13 residues that are also responsible for CsA binding (Fig. 1A), is identical to that of Cyp3, CypA, and TaCypA-1 (Pflügl et al, 1993;Ke et al, 1994;Dornan et al, 1999;Sekhon et al, 2013). Structure alignment of CsCyp with TaCypA-1, the only other plant divergent Cyp with known threedimensional (3D) structure, showed no major structural differences (root mean square deviation = 0.43 Å).…”

Section: On the Structural (Dis)similarities Between Classical And DImentioning

confidence: 92%

A Redox 2-Cys Mechanism Regulates the Catalytic Activity of Divergent Cyclophilins

et al. 2013

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The citrus (Citrus sinensis) cyclophilin CsCyp is a target of the Xanthomonas citri transcription activator-like effector PthA, required to elicit cankers on citrus. CsCyp binds the citrus thioredoxin CsTdx and the carboxyl-terminal domain of RNA polymerase II and is a divergent cyclophilin that carries the additional loop KSGKPLH, invariable cysteine (Cys) residues Cys-40 and Cys-168, and the conserved glutamate (Glu) Glu-83. Despite the suggested roles in ATP and metal binding, the functions of these unique structural elements remain unknown. Here, we show that the conserved Cys residues form a disulfide bond that inactivates the enzyme, whereas Glu-83, which belongs to the catalytic loop and is also critical for enzyme activity, is anchored to the divergent loop to maintain the active site open. In addition, we demonstrate that Cys-40 and Cys-168 are required for the interaction with CsTdx and that CsCyp binds the citrus carboxyl-terminal domain of RNA polymerase II YSPSAP repeat. Our data support a model where formation of the Cys-40-Cys-168 disulfide bond induces a conformational change that disrupts the interaction of the divergent and catalytic loops, via Glu-83, causing the active site to close. This suggests a new type of allosteric regulation in divergent cyclophilins, involving disulfide bond formation and a loop-displacement mechanism.

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Crystal structures of peptides and modified peptides

Marraud

Aubry

1996

Biopolymers

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The X‐ray diffraction experiments on peptides and related molecules which have been carried out in Western Europe, except Italy, in the last eight years are reviewed. The crystal structures of some bioactive peptides such as Leu‐enkephalin (a neurotransmitter), cyclosporin A (an immunomodulator in both the free and protein‐bound state), balhimycin (an antibiotic) and octreotide (a somatostatin analogue) are briefly presented. Crystallized N‐ and C‐protected model peptides have given an insight into the folding tendency and folding modes depending on the peptide sequences. The crystal structures of various pseudopeptide molecules reveal how the three‐dimensional structure of peptide analogues can be modulated by substituting non‐peptide groups for the peptide bond. A few examples of structural mimetics of the β‐ and γ‐turns, and of templates for α‐helix induction are also presented. © 1996 John Wiley & Sons, Inc.

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Crystal structures of cyclophilin A complexed with cyclosporin A and N-methyl-4-[(E)-2-butenyl]-4,4-dimethylthreonine cyclosporin A

Cited by 83 publications

References 50 publications

Crystal structure of calcineurin–cyclophilin–cyclosporin shows common but distinct recognition of immunophilin–drug complexes

Crystal structure of calcineurin–cyclophilin–cyclosporin shows common but distinct recognition of immunophilin–drug complexes

A Redox 2-Cys Mechanism Regulates the Catalytic Activity of Divergent Cyclophilins

Crystal structures of peptides and modified peptides

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