Pyrimidine Dimer Class II Photolyase in Complex with UV
Kelsey Dillon '14 and Jenny Shoots '14
UV light can damage DNA
by causing the formation of pyrimidine dimers. These dimers distort the
DNA backbone, impairing replication and transcription. Photolyases are
part of the DNA repair machinery of archaea, bacteria, and plants, but
are not found in placental mammals. They function by breaking the
cyclobutane ring of pyrimidine dimers. Photolyases function only in the
presence of blue, near-UV light. They contain at least one chromophore
cofactor to harvest energy from light to catalyze repair.
structural study of photolyases to date have been of class I
photolyases, found exclusively in microbes. The below tutorial is about
the first crystal structure of a class II photolyase. It was isolated
from the archaea Methanosarcina mazei,
but multicellular eukaryotes
possess class II photolyases as well. The class I, II, and III
divisions were created based on sequence differences. This tutorial
outlines a few of the structural differences between class I and class
II photolyases. The chromophore cofactor of this photolyase, referred
to as MmCPII•CPD, is FAD. Functionally, it is photoexcited to
form. It can then transfer an electron into the cyclobutane
pyrimidine dimer (CPD), breaking the two carbon-carbon bonds linking
the bases together.
structure of MmCPDII is organized in an N-terminal subdomain
of α helices and β sheets.
subdomain is comprised of
only alpha helices.
two subdomains are
joined by a linker.
linker is very
flexible, and much of its intervening structure is unknown.
the photolyase is
complex with UV-damaged DNA, it is the C-terminal
which contacts the thymidine
cofactor FAD also
makes its contacts with the C-terminal
FAD Binding Site
require a chromophore cofactor, in this case FAD, for both recogniction
of CPD lesions and for catalysis. Upon
excitation by light, photolyases reduce their cofactor FAD into FADH-.
This activity requires
a photoreduction pathway which transfers excited electrons to FAD,
shown here in its oxidized form.
depend on three linear tryptophan
residues for electron transfer. The class II photolyase shown here
several tryptophan and tyrosine residues which can transfer electrons
but photoreduction has
been shown to be dependent only upon two tryptophans.
transferred from W360
residue N403 stabilizes FAD during photoreduction.
Interactions with Pyrimidine Dimer
the MmCPDII•CPD-DNA complex, the DNA has a synthetic CPD
lesion in its central
position. The dsDNA adopts a typical B-type
conformation, but is severely
at the lesion
site. The CPD lesion
of the kinked dsDNA is flipped out
of the duplex into the active site
of MmCPDII and bound there.
MmCPDII, conserved tryptophans W305
form the L-shaped walling
of the active site that clamps the CPD lesion together with the side
chain of the
repair a transient radical form of the CPD lesion forms once
FADH- transfers an electron to the lesion. E301
stabilizes the CPD radical anion by proton transfer after the breaking
of the thymine dimer. It forms
hydrogen bonds with the 5’-thymine of the lesion.
cluster of six water molecules
the space between the CPD lesion
and the active site’s walling.
of these water
molecules replaces the
conserved side chain asparagine found in class I
photolyases (replaced by a glycine in MmCPDII) by taking over the
bridging interaction between the C4-carbonyl group of the
3’-thymine and the C2’-hydroxl of the
FAD’s ribityl group.
other five water molecules
located as a square-pyrimidal cluster between the 3’-thymine
and the diphosphate group of FAD.
Kim, S.-T. and A. Sancar
1993. Photochemistry, Photophysics, and
Mechanism of Pyrimidine Dimer Repair by DNA Photolyase. Photochemistry
and Photobiology, 57: 895–904.
Kiotntke, S., Geisselbrecht,
R. Carell, T., Batschaer, A., and L.-O. Essen. 2011. Crystal structures
of an archael class II DNA photolyase and its complex with UV-damaged
duplex DNA. EMBO Jour. 30: 4437-4449.
Malhotra, K., Kim, S.-T.,
Walsh, C., and A. Sancar. 1992. Roles of FAD and
8-Hydroxy-5-deazaflavin Chromophores in Photoreactivation of by Anacystis
nidulans DNA photolyase. J. Biol
Chem. 267(22): 15406-15411.