Table of Contents:
Cryptochromes and photolyases constitute a group of highly distributed, FAD-binding,
bluelight dependent signaling proteins ans enzymes, which together form the
photolyase/cryptochrome-superfamily (PCSf). Photolyases recognize UV-induced DNA
lesions between adjacent pyrimidine bases, namely cyclobutanpyrimidindimers (CPD) and
(6-4)-pyrimidine-pyrimidone-photoproducts ((6-4)), and repair them blue-light dependently,
whereas cryptochromes rather show regulatory functions in vivo. The plant representatives are
involved in responses to blue-light stimuli and take influence on growth, development and the
circadian clock of the plant. Cryptochromes in animals participate in the circadian clock as
blue-light sensors (type I), or light-independently as part of the central oscillator (type II).
Cryptochromes from bacteria are still poorly understood, the best characterized
representatives from Synechocystis sp. PCC6803 belongs to the CryDASH family whose
exact biological function is still unknown.
This work dealt with functional and the first structural characterization of a true bacterial
cryptochrome,cryptochrome B from Rhodobacter sphaeroides (RsCryB). RsCryB shows no
DNA repair activity and participates in the light-dependent and oxygen-dependent regulation
of photosynthesis genes in R. sphaeroides. It defines a new family of proteins in the PCSf,
predominantly occurring in proteobacteria, the proteobacterial cryptochromes (CryPro).
Despite low sequence identity to the other representatives of the PCSf, the structure of the
CryPro family is analogous to the conserved overall fold of the superfamily. Surprisingly, one
[4Fe-4S] clusters was identified, which is along with the catalytic cofactor FAD the defining
element of RsCryB's C-terminus. This cluster is structurally and chemically related to known
clusters of eukaryotic primase subunits, as was shown by EPR experiments. Moreover 6,7-
dimethyl-8-ribityl-lumazine was identified as antenna chromophore of RsCryB, yet unknown
inside the PCSf.
These studies are complemented by an DNA binding analysis of the class II CPD photolyase
from Methanosarcina mazei (MmCPDII), the model photolyase for plant homologs, as well as
an analysis of the fully reduced state of MmCPDII by ultrafast spectroscopy. In the third part
of the project the native antenna chromophore of the (6-4)-photolyase from Dunaliella salina
was identified as 8-hydroxy-5-deazaflavin and the in vitro repair of (6-4)-damages by the
enzyme was shown.