shifts predictors from structural ensembles have been developed
[91–94].
It is important to emphasize that unbiased MD simulations of
some hundreds of nanoseconds or even microseconds are unlikely
to provide enough statistical power to be used to sample conformational changes that are related to long timescale dynamics like
the one revealed by the p53 NMR experiments. Indeed, in classical
MD even using multiple replicate approaches, only a few transitions
could be observed among different states, whereas a proper investigation would require the sampling for multiple times of the same
event. When we simulated p53 DBD without the N-terminal tail,
we observed higher flexibility of the S6-S7 loop in the ns timescale.
This observation raises serious concerns for the usage of a DBD
construct lacking the N-terminal disordered residues to study the
properties of the DBD regions that can be modulated by intramolecular interaction with the tail [14]. Indeed, such fast motions
were not expected. Supporting this notion, classical unbiased MD
simulations of the construct including the tail (91–289) did not
show any substantial differences between the DNA-bound and
-unbound forms of the loop [14], indicating that different techniques for conformational sampling needed to be applied.
In our study [14], we applied the framework above to the study
of p53 DBD in its unbound, DNA-bound, and phosphorylated
state and on a specific region of the protein (S6-S7 loop). The work
can be envisaged as a proof of concept for future applications to
unveil the complexity of the p53 signaling function. Indeed, the
novelty of our work is not so much about the fact that we identified
a coupling between conformational rearrangements at the interface
for DNA-binding and changes in a loop (S6-S7 loop, residues
207–213). Other previous works already suggested a long-range
coupling in the proximity of the S6-S7 loop using classical MD only
and techniques such as principal component analyses [39]. More
importantly, we showed that the conformational changes in the L1
loop at the DNA-binding interface are tightly coupled to changes
in the S6-S7 loop, which in turn is in proximity to the N-terminal
disordered tail, which is also involved in the mechanism (Fig. 2).
We showed with high accuracy that DNA modulates the conformational ensemble of the S6-S7 loop conformational ensemble. We
also identified key residues that are involved in the paths of structural communication between the two distal sites and that also
include the N-terminal disordered region. The proposed mechanism still holds in the context of the p53 tetramer, as shown by a
comparison of the structure samples by the p53 DBD simulations
with the known experimental structures of the p53 quaternary
assembly [14]. Indeed, the different S6-S7 conformations fit into
the tetramer without clashes and with most of the residues solved
exposed and available for interaction in S6-S7 more “open” states.
Dynamics of p53
229
[91–94].
It is important to emphasize that unbiased MD simulations of
some hundreds of nanoseconds or even microseconds are unlikely
to provide enough statistical power to be used to sample conformational changes that are related to long timescale dynamics like
the one revealed by the p53 NMR experiments. Indeed, in classical
MD even using multiple replicate approaches, only a few transitions
could be observed among different states, whereas a proper investigation would require the sampling for multiple times of the same
event. When we simulated p53 DBD without the N-terminal tail,
we observed higher flexibility of the S6-S7 loop in the ns timescale.
This observation raises serious concerns for the usage of a DBD
construct lacking the N-terminal disordered residues to study the
properties of the DBD regions that can be modulated by intramolecular interaction with the tail [14]. Indeed, such fast motions
were not expected. Supporting this notion, classical unbiased MD
simulations of the construct including the tail (91–289) did not
show any substantial differences between the DNA-bound and
-unbound forms of the loop [14], indicating that different techniques for conformational sampling needed to be applied.
In our study [14], we applied the framework above to the study
of p53 DBD in its unbound, DNA-bound, and phosphorylated
state and on a specific region of the protein (S6-S7 loop). The work
can be envisaged as a proof of concept for future applications to
unveil the complexity of the p53 signaling function. Indeed, the
novelty of our work is not so much about the fact that we identified
a coupling between conformational rearrangements at the interface
for DNA-binding and changes in a loop (S6-S7 loop, residues
207–213). Other previous works already suggested a long-range
coupling in the proximity of the S6-S7 loop using classical MD only
and techniques such as principal component analyses [39]. More
importantly, we showed that the conformational changes in the L1
loop at the DNA-binding interface are tightly coupled to changes
in the S6-S7 loop, which in turn is in proximity to the N-terminal
disordered tail, which is also involved in the mechanism (Fig. 2).
We showed with high accuracy that DNA modulates the conformational ensemble of the S6-S7 loop conformational ensemble. We
also identified key residues that are involved in the paths of structural communication between the two distal sites and that also
include the N-terminal disordered region. The proposed mechanism still holds in the context of the p53 tetramer, as shown by a
comparison of the structure samples by the p53 DBD simulations
with the known experimental structures of the p53 quaternary
assembly [14]. Indeed, the different S6-S7 conformations fit into
the tetramer without clashes and with most of the residues solved
exposed and available for interaction in S6-S7 more “open” states.
Dynamics of p53
229