Thanks to the recent development of force field parametrization, MD simulations
are currently quite reliable. It is still quite common that results from the force field
methods are not fully in line with empirical data, and this is especially true if we
consider X-ray crystallography. Long timescale simulation studies with p38a MAP
kinase inhibitors were recently used to explain the discrepancy between X-ray and
NMR data [50]. According to the classical activation mechanism, supported by
X-ray structures [51], p38a MAP kinase activation with a double-phosphorylated
structure should include a large reorientation of the activation loop A. However,
NMR studies indicated that double phosphorylation does not induce any major
conformational rearrangements [52]. Simulations with CHARMM force field were
conducted with ten replicas, and simulation time was varied between 500 ns and
1 μs, although in individual cases also longer simulation times were used. The results
suggest that p38a predominantly samples conformations which are in contrast with
the activation model obtained from X-ray crystallography. However, the authors
analyzed crystal contacts and found several artifacts affecting the protein conformation, for example, an atypically long expression His-tag of a neighboring molecule
bound to the hydrophobic docking groove of p38a MAP kinase. It is easy to agree
with the statement of the Kuzmanic et al. [50] “These observations show how
important it is to carefully analyze symmetry-related molecules and they call for
caution in the interpretation of deposited X-ray structures, as they can be
misleading.”
Basically, the abovementioned conclusion can be drawn also from the studies
dealing with Aurora kinase A (AurA) [53, 54]. By combining experimental data and
MD simulations, it has been demonstrated that AurA activation by phosphorylation
occurs without a population shift from the DFG-out to the DFG-in state and that the
activation loop of the activated kinase remains highly dynamic. This is, once more,
against the traditional view of the X-ray. Instead, molecular dynamics simulations
and electron paramagnetic resonance experiments show that phosphorylation triggers a switch within the DFG-in subpopulation from an autoinhibited DFG-in
substate to an active DFG-in substate, leading to catalytic activation.
4 Allosteric Control of Kinases
Most of the kinase inhibitors target the ATP binding site of the corresponding kinase
protein. While the ATP binding site is highly conserved among the kinome, the
so-called exosites are much more unique, although to some extent also conserved.
The first successful kinase inhibitor targeting exosites was imatinib [55]. From the
modeling point of view, this paradigm shift was quite big as it demonstrated that
target protein conformation is not static and that kinase conformation can be
modified by targeting exosites. While, at the moment, most of the new kinase
inhibitors are targeting the ATP-binding cleft between the N- and C-lobes of the
kinase, interest toward allosteric inhibitors is growing due to some very evident
benefits. The biggest advantage is the fact that the allosteric binding site has no
34
A. Poso
are currently quite reliable. It is still quite common that results from the force field
methods are not fully in line with empirical data, and this is especially true if we
consider X-ray crystallography. Long timescale simulation studies with p38a MAP
kinase inhibitors were recently used to explain the discrepancy between X-ray and
NMR data [50]. According to the classical activation mechanism, supported by
X-ray structures [51], p38a MAP kinase activation with a double-phosphorylated
structure should include a large reorientation of the activation loop A. However,
NMR studies indicated that double phosphorylation does not induce any major
conformational rearrangements [52]. Simulations with CHARMM force field were
conducted with ten replicas, and simulation time was varied between 500 ns and
1 μs, although in individual cases also longer simulation times were used. The results
suggest that p38a predominantly samples conformations which are in contrast with
the activation model obtained from X-ray crystallography. However, the authors
analyzed crystal contacts and found several artifacts affecting the protein conformation, for example, an atypically long expression His-tag of a neighboring molecule
bound to the hydrophobic docking groove of p38a MAP kinase. It is easy to agree
with the statement of the Kuzmanic et al. [50] “These observations show how
important it is to carefully analyze symmetry-related molecules and they call for
caution in the interpretation of deposited X-ray structures, as they can be
misleading.”
Basically, the abovementioned conclusion can be drawn also from the studies
dealing with Aurora kinase A (AurA) [53, 54]. By combining experimental data and
MD simulations, it has been demonstrated that AurA activation by phosphorylation
occurs without a population shift from the DFG-out to the DFG-in state and that the
activation loop of the activated kinase remains highly dynamic. This is, once more,
against the traditional view of the X-ray. Instead, molecular dynamics simulations
and electron paramagnetic resonance experiments show that phosphorylation triggers a switch within the DFG-in subpopulation from an autoinhibited DFG-in
substate to an active DFG-in substate, leading to catalytic activation.
4 Allosteric Control of Kinases
Most of the kinase inhibitors target the ATP binding site of the corresponding kinase
protein. While the ATP binding site is highly conserved among the kinome, the
so-called exosites are much more unique, although to some extent also conserved.
The first successful kinase inhibitor targeting exosites was imatinib [55]. From the
modeling point of view, this paradigm shift was quite big as it demonstrated that
target protein conformation is not static and that kinase conformation can be
modified by targeting exosites. While, at the moment, most of the new kinase
inhibitors are targeting the ATP-binding cleft between the N- and C-lobes of the
kinase, interest toward allosteric inhibitors is growing due to some very evident
benefits. The biggest advantage is the fact that the allosteric binding site has no
34
A. Poso