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Today, I will review photoactivation mechanism
of cryptochrome2. In the last presentation, I reviewed photo-induced
protein-protein interaction between CRY2 and CIBN. The basic working principle was simple.
CRY2 undergoes conformational change catalyzed by blue light, and then N-terminal photolyase
homology region (PHR) of CRY2 reversibly associating with the CIBN. In this context, one question
is hit upon. How can CRY2 be changed and bind to CIB1 by blue light? There is one hypothesis,
that is, tryptophan-triad dependent photoreduction.
To understand the detail mechanism, I will
preferentially review the family of crytochromes and its structure. Here is a phylogenetic
tree of cryptochromes. There are various cryptochromes in Nature. Green indicates plant cryptochromes,
pink and violet indicate animal, blue are in DASH, and red brown gray orange color represent
photolyases. In the course of evolution, major cryptochromes have arisen from separate photolyase
ancestors. So whether, the crystal structure of cryptochromes and photolyases are similar,
however, the functions are totally different.
Cryptochromes are originated in plants and
animals, those are signaling molecules which regulate diverse biological responses such
as entrainment of circadian rhythms in plants and animals. In contrary, photolyases are
enzymes in prokaryotics which can utilize light energy for repair of UV damaged DNA.
On the other hand, cryptochromes and photolyases have high similarity and identity, and also
have similar photoactive domain which is called Flavin adenin dinucletides assess site. This
site is a key point for photo-induced conformational change of the proteins.
A proposed photocycle of cryptochrome whereby
light-induced radical accumulation occurs subsequent to illumination, to be followed
by reoxidation upon return to darkness, is shown in Figure. When the blue light irradiate
FAD in CRY, an anionic flavin radical FAD forms the resting state and that direct electron
transfer from flavin to substrate is the basis for signaling, or FAD receives hydrogen ion
and forms to FADH that direct leads to conformational change of CRY strucuture. The important feature
to note is that cry, unlike photolyases, undergoes reversible cycling between primarily oxidized
and radical flavin states subsequent to illumination.
The main characters that change of FAD are
tryptophan-triad. The electron transfer chain that reduces photoexcited FAD and FADH◦
via a conserved tryptophan-triad. Cryptochrome photoactivation is triggered by blue-light
photoexcitation of the FAD cofactor initially present in the oxidized state. Excited flavin,
FAD*, receives an electron from one of the nearby tryptophans. Electron transfer from
tryptophan leads to formation of an ionic FAD•− + W(H)•+ radical pair, which is
then transformed into a stable neutral FADH• + W• radical pair state through proton exchange
with the nearby other W. Thus, the structure of CRY is changed by produced FADH. After
reaction, under no blue light, the neutral radical pair recombines back to the initial
state through coupled electron−proton transfer.
In case of CRY2 in Arabidopsis thaliana, W321,
375, and 397 are main molecules of tryptophan-triad dependent photoreduction. Indeed, a mutation
at any of the trp-triad residues of CRY2 effectively reduced or abolished the photoreduction activity.
Until now, we has revealed how CRY2 is changed
by the blue light. Then, how can CRY2 bind to CIBN by the blue light? Unfortunately,
the mechanism has not discovered yet. To prove the concept, we should consider three
factors at least. First, primary light-driven photoreaction. Analysis of a light-driven
reaction of cryptochromes that causes productive interaction with a biologically relevant substrate
in vitro is required. Second, the structure of activated form of cryptochrome. Cocrystalization
with a signaling partner is required. At last, magnetoreception. The identity of the biologically
relevant magnetosensitive reaction (radical pair) needs to be established for cryptochromes.
This discover will provide the better understanding of cell signaling and its controllable engineering
field based on light-induced protein-protein interaction.