Updated structure of Drosophila cryptochrome

Abstract

We report an improved model of the Drosophila cryptochrome structure that corrects errors in the original coordinates (3TVS.pdb). Further refinement of the structure with automated rebuilding algorithms in PHENIX 1 followed by manual building, indicated that a model of dCRY could be produced with excellent refinement statistics without taking into account the non-merohedral twinning originally reported (Table 1). The rebuilt structure has an RMSD on Cα positions of 2.4 Å compared to the deposited coordinates with most differences found in the conformation of surface loops (Fig. 1). However, the new analysis also indicates that the sequence register of the C-terminal tail helix (CTT) is displaced by two residues (Fig. 2). This change in sequence register offsets the invariant FFW motif along the helix axis such that Phe534, and not Trp536, approaches closest to the flavin ring (Fig. 3). In the new model, the three residues composing the FFW motif continue to make extensive interactions with the photolyase homology domain. This new position of the FFW motif is more consistent with the cellular data of Fig. S6, which shows that substitution of FFW to three alanine residues has a dramatic effect on dCRY stability, but that the W536A substitution alone, does not. The configuration of the flavin center is similar between the old and new models, with the largest difference in the angle of the ribityl-to-flavin (N10) bond (Fig. 4). Phosphorylation of Thr518 is not apparent in the new electron density maps despite identification of this modification by mass spectrometry. The errors in the original structure stemmed from model bias introduced during the detwinning procedure. Lower resolution data sets to which the original dCRY structure was built appear to suffer more from twinning than the 2.3 Å resolution data that the final model was refined against. Although the high-resolution data does contain indications of non-merohedral twinning, including intensity oscillations along the reciprocal space l axis and spurious Patterson peaks, a model that agrees well with the diffraction data as collected can be produced without compensation for these effects (Table 1). The new coordinates have been deposited in the PDB as 4GU5. B.R.C. apologizes for these errors.

Phosphorylation of the Cryptochrome 1 C-terminal Tail Regulates Circadian Period Length

“…This model is supported by recent crystal structures, which show that the C-terminal helix docks in a groove of the PHR domain that is analogous to the DNA-binding groove in photolyase (13)(14)(15). Mammalian CRYs are not activated by light, and their C-terminal tails are not conserved with the Drosophila CRY C-terminal tail.…”

Flavin reduction activates Drosophila cryptochrome

“…The sequence register of the CTT in the original dCRY structure was incorrect by two residues but has since been updated in an improved model (8) that also agrees with a more recent structure of dCRY (31). The CTT is well poised to respond to light-induced redox changes at the FAD.…”

“…The conserved FFW motif of type 1 CRYs (F534, F535, and W536) associates closely with the PHD by binding into the flavin pocket (7,8) (Fig. 2A).…”

“…CRYs are closely related to the photolyase (PL) family of proteins that repair UV-damaged DNA in a blue light-dependent fashion (1,3,5). All PLs and CRYs have a FAD chromophore bound in an α-helical domain, and many also have a pteridine or deazaflavin antenna cofactor bound in an α/β-nucleotide-binding domain (5)(6)(7)(8). Type I insect CRYs do not bind an antenna cofactor (7,9).…”

The quantum needle of the avian magnetic compass

“…A magnetic field strength of 50 μT was used throughout. The relative orientation of the two radicals was that of FAD and Trp-342 in Drosophila melanogaster cryptochrome (Protein Data Bank ID code 4GU5) (SI Appendix, Section S1) (42,43). The initial state of the spin system was a pure singlet.…”