Structural changes within the bifunctional cryptochrome/photolyase CraCRY upon blue light excitation

Area of Science:

• Biochemistry
• Molecular Biology
• Chronobiology

Background:

• Cryptochromes (CRYs) are blue-light photoreceptors crucial for circadian rhythms in animals, sharing ancestry with DNA-repairing photolyases (PHLs).
• A key structural difference is the variable C-terminal extension (CTE) in CRYs, whose function remains largely speculative.
• The green algae Chlamydomonas reinhardtii possesses an animal-like cryptochrome (CraCRY) with potential bifunctional roles.

Purpose of the Study:

• To investigate the structural role and binding interactions of the CTE in Chlamydomonas reinhardtii cryptochrome (CraCRY).
• To explore the light-dependent conformational changes of CraCRY using advanced biophysical techniques.
• To develop a method for analyzing photoreceptors in defined photostates.

Main Methods:

• Hydrogen/deuterium exchange coupled with mass spectrometry (HDX-MS) to map protein dynamics.
• Development of a 'light chamber' for automated HDX-MS measurements under controlled light conditions.
• Comparison of oxidized and reduced flavoprotein states of CraCRY.

Main Results:

• The CTE of CraCRY was found to bind to the surface of the photolyase homology region, near the DNA binding site.
• Significant differences in protein dynamics were observed between the oxidized and reduced flavoprotein states.
• A model for light-dependent switching of CraCRY's bifunctional activity was proposed based on these findings.

Conclusions:

• The CTE plays a structural role in modulating the activity of cryptochromes.
• Light induces conformational changes in CraCRY, suggesting a mechanism for light-dependent regulation.
• This study provides insights into the evolution and function of cryptochromes as bifunctional proteins.