Chemistry Reference
In-Depth Information
The main dimeric sites are residues Asp260 to Val263 at the N-terminus of an -helix (262-273) from one
monomer, which interacts with Tyr216 to Arg220 at the end of the activation segment of the other. Direct polar
interactions are limited to an ion pair between Asp260 and Arg220, and main chain to main chain hydrogen bonds
between the peptide carbonyls of Asp260 and Val263 with the peptide nitrogens of Arg220 and Tyr216,
respectively. The hydrophobic face of the side chain of Tyr216 interacts with a hydrophobic patch formed by Ile228,
Phe229, Gly262, Val263, and Leu266 from the other monomer. There are overall shape complementarities at the
interface but few other direct contacts. Several solvent-bridged interactions are evident. In general, dimer formation
buries ~2358 Å
2
of accessible surface [59].
The overall shape is shared by all kinases [61,62], consisting of an N-terminal β-strand domain (residues 25-138)
and a C-terminal α-helical domain (residues 139-343) [60]. The N-terminal domain is formed by a seven-stranded
sheet curved over on itself to form a closed orthogonal barrel. The 5th and 6th strands of the barrel are connected
by a short -helix (residues 96-102) that consists of two turns. This helix is conserved in all kinases, whereas two of
its residues play key roles in the catalytic activity of the enzyme. Arg96 is involved in the alignment of the two
domains. Glu97 is positioned in the active site and forms a salt bridge with Lys85, a key residue in catalysis. The N-
terminal domain connects to the remainder of the protein via an -helix (residues 138-149) extending from the end
of the 7th strand [60].
The activation loop (residues 200-226) runs along the surface of the substrate binding groove. The C-terminal 39
residues (residues 344-382) are outside the core kinase fold and forms a small domain that packs against the -
helical domain. The ATP (adenosine triphosphate) binding site (substrate-binding pocket) is at the interface of the
-helical and -strand domain and is bordered by the glycine-rich loop and the hinge. It contains three basic
residues, i.e. Arg96, Arg180 and Lys205, that bind the phosphate anion on the primed Ser/Thr residue in the
substrate motif [59,60].
CATALYTIC ACTIVITY AND REGULATION OF GSK-3
The catalytic activity of protein kinases depends upon the correct orientation of the catalytic groups contributing to
the transfer of the -phosphate group from ATP to a Ser, Thr, or Tyr side chain of the protein substrate. Another
factor is the accessibility and correct positioning of the groups forming the substrate peptide binding site, which
provides affinity and specificity for the substrate [59,63].
GSK3- has two phosphorylation sites, that influence the catalytic activity of the protein. Ser9 is the
phosphorylation site for AKT, and the phosphorylation of this residue inactivates GSK3-. Phosphorylation of
Tyr216, located on the activation loop, increases the catalytic activity.
The substrate specificity of GSK3β is unusual. Recognition of substrates by the GSK-3 usually requires prior
phosphorylation of the substrate. Analysis of phosphorylated sites by GSK-3 [59] (see Table
2
) suggests a
preference for sequences of the type: Ser/Thr-X-X-X-Ser/Thr, where the C-terminal serine or threonine is already
phosphorylated [64-66], so that prior phosphorylation of the n+4th residue facilitates phosphorylation of the nth
residue.
GSK-3 function can be regulated at many levels [38,67]. GSK-3 can be negatively regulated by phosphorylation
at the N-terminal domain (Ser9 residue), which acts as a pseudo-substrate that blocks the access of substrates to the
catalytic site [68] or can be activated through phosphorylation of Tyr216 in the T-loop domain, which facilitates the
access of substrates to the catalytic site [69]. In addition, the activity of this protein can be controlled through its
intracellular distribution [70], and by its interaction with many other proteins [71] or potential inhibitors [69].
GSK-3
INHIBITORS
Regarding the high therapeutical potential of targeting GSK-3β in many different pathologies, the search for its
inhibitors is a very active field in both academic centers and pharmaceutical companies. For approximately the last
fifty years the drug lithium has been the mainstay for the treatment of bipolar disorder, with a beneficial effect often
observed in approximately 60-80% of patients and with no tolerance or sensitivity been developed during many
