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Molecular assembly of the period-cryptochrome circadian transcriptional repressor complex.

Nangle SN, Rosensweig C, Koike N, Tei H, Takahashi JS, Green CB, Zheng N - Elife (2014)

Bottom Line: PER2-CBD adopts a highly extended conformation, embracing CRY2 with a sinuous binding mode.Its N-terminal end tucks into CRY adjacent to a large pocket critical for CLOCK-BMAL1 binding, while its C-terminal half flanks the CRY2 C-terminal helix and sterically hinders the recognition of CRY2 by the FBXL3 ubiquitin ligase.Unexpectedly, a strictly conserved intermolecular zinc finger, whose integrity is important for clock rhythmicity, further stabilizes the complex.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, University of Washington, Seattle, United States.

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Related in: MedlinePlus

Major differences between CRY1-PER2-CBD and CRY2-PER2-CBD complex structures.Superposition of the two structures reveals major structural dissimilarities between the two paralogs at the CRY secondary pocket and a residual fusion-protein sequence (yellow) in CRY1-bound PER2-CBD. The PER2-CBD (dark blue) N-terminus together with the artifactual sequence (AGLEVLFQGPDSM) forms a β-hairpin and induces an inward conformation of the CRY1 (light green) serine loop.DOI:http://dx.doi.org/10.7554/eLife.03674.014
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fig4s3: Major differences between CRY1-PER2-CBD and CRY2-PER2-CBD complex structures.Superposition of the two structures reveals major structural dissimilarities between the two paralogs at the CRY secondary pocket and a residual fusion-protein sequence (yellow) in CRY1-bound PER2-CBD. The PER2-CBD (dark blue) N-terminus together with the artifactual sequence (AGLEVLFQGPDSM) forms a β-hairpin and induces an inward conformation of the CRY1 (light green) serine loop.DOI:http://dx.doi.org/10.7554/eLife.03674.014

Mentions: During the preparation of this manuscript, the complex structure of mammalian CRY1-PHR and PER2-CBD was reported (Schmalen et al., 2014). With high sequence conservation between CRY1 and CRY2, PER2-CBD adopts a similar CRY-binding mode with a tetrahedral coordination of a zinc ion by an intermolecular CCCH zinc-binding motif. The major structural difference lies at the interface of the N-terminal region of PER2-CBD and the CRY secondary pocket. The CRY1-bound PER2-CBD fragment contains a residual fusion-protein sequence, which forms an artifactual ß-hairpin with the first five amino acids of the PER2-CBD (Figure 4—figure supplement 3). In contrast to the PER2-bound CRY2 serine loop, but reminiscent of the Drosophila CRY antenna loop (Zoltowski et al., 2011), the otherwise disordered (Czarna et al., 2013) CRY1 serine loop adopts an inward conformation and occludes the secondary pocket. This conformational difference reveals a substantial degree of structural plasticity, which might be necessary for differential binding and regulation at this site. Interestingly, Schmalen et al. (2014) identified a potential redox sensor involving a disulfide bond near the zinc finger between Cys412 and Cys363, which modulates CRY1-PER2 binding. However, in our circadian reporter assay, we did not detect any difference between the CRY1 wild type and C412A mutant (Figure 5D). More in-depth analyses can now exploit the specific structural differences between the two complexes to explain the non-redundant roles of the two Cryptochrome proteins.


Molecular assembly of the period-cryptochrome circadian transcriptional repressor complex.

Nangle SN, Rosensweig C, Koike N, Tei H, Takahashi JS, Green CB, Zheng N - Elife (2014)

Major differences between CRY1-PER2-CBD and CRY2-PER2-CBD complex structures.Superposition of the two structures reveals major structural dissimilarities between the two paralogs at the CRY secondary pocket and a residual fusion-protein sequence (yellow) in CRY1-bound PER2-CBD. The PER2-CBD (dark blue) N-terminus together with the artifactual sequence (AGLEVLFQGPDSM) forms a β-hairpin and induces an inward conformation of the CRY1 (light green) serine loop.DOI:http://dx.doi.org/10.7554/eLife.03674.014
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4157330&req=5

fig4s3: Major differences between CRY1-PER2-CBD and CRY2-PER2-CBD complex structures.Superposition of the two structures reveals major structural dissimilarities between the two paralogs at the CRY secondary pocket and a residual fusion-protein sequence (yellow) in CRY1-bound PER2-CBD. The PER2-CBD (dark blue) N-terminus together with the artifactual sequence (AGLEVLFQGPDSM) forms a β-hairpin and induces an inward conformation of the CRY1 (light green) serine loop.DOI:http://dx.doi.org/10.7554/eLife.03674.014
Mentions: During the preparation of this manuscript, the complex structure of mammalian CRY1-PHR and PER2-CBD was reported (Schmalen et al., 2014). With high sequence conservation between CRY1 and CRY2, PER2-CBD adopts a similar CRY-binding mode with a tetrahedral coordination of a zinc ion by an intermolecular CCCH zinc-binding motif. The major structural difference lies at the interface of the N-terminal region of PER2-CBD and the CRY secondary pocket. The CRY1-bound PER2-CBD fragment contains a residual fusion-protein sequence, which forms an artifactual ß-hairpin with the first five amino acids of the PER2-CBD (Figure 4—figure supplement 3). In contrast to the PER2-bound CRY2 serine loop, but reminiscent of the Drosophila CRY antenna loop (Zoltowski et al., 2011), the otherwise disordered (Czarna et al., 2013) CRY1 serine loop adopts an inward conformation and occludes the secondary pocket. This conformational difference reveals a substantial degree of structural plasticity, which might be necessary for differential binding and regulation at this site. Interestingly, Schmalen et al. (2014) identified a potential redox sensor involving a disulfide bond near the zinc finger between Cys412 and Cys363, which modulates CRY1-PER2 binding. However, in our circadian reporter assay, we did not detect any difference between the CRY1 wild type and C412A mutant (Figure 5D). More in-depth analyses can now exploit the specific structural differences between the two complexes to explain the non-redundant roles of the two Cryptochrome proteins.

Bottom Line: PER2-CBD adopts a highly extended conformation, embracing CRY2 with a sinuous binding mode.Its N-terminal end tucks into CRY adjacent to a large pocket critical for CLOCK-BMAL1 binding, while its C-terminal half flanks the CRY2 C-terminal helix and sterically hinders the recognition of CRY2 by the FBXL3 ubiquitin ligase.Unexpectedly, a strictly conserved intermolecular zinc finger, whose integrity is important for clock rhythmicity, further stabilizes the complex.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, University of Washington, Seattle, United States.

Show MeSH
Related in: MedlinePlus