Ligand
Docking to Biochemical Targets. Crystallography and modelling
J. Hašek, J. Dohnálek, J. Dušková, P. Kolenko, T. Skálová
Institute
of Macromolecular Chemistry,
hasek@imc.cas.cz
Keywords:
Ligand-protein docking, polymer, drug design, X-ray structure analysis
X-ray structure analysis is an excellent tool
for study of molecular recognition (i.e. specific adhesion between two
macromolecules of biological origin) and therefore it plays a key role in
elucidation of molecular mechanisms of many biochemical processes. A high
interest of crystallographers was also devoted to specific interactions of
drugs in active sites of enzymes and their research has already been reflected
in many practical results of the rational drug design. Also in the case of
drugs, we can usually see high affinity of the ligand to the target protein
because it is the aim of the human effort to break down the enzymatic function
permanently. The molecules of interest are in these cases usually well ordered
in crystal and therefore relatively easily resolved by protein crystallography.
The situation is not so easy with ligands
possessing lower affinity to the protein molecule (as for example polymers).
One has to cope usually with molecules
which are only partly localized on the protein surface with the remaining parts
floating freely in the crystallization buffer (for diffraction experiment
invisible). In addition, parts of the ligand adhering to protein surface (the
only visible fragments in the maps of electron density) have often lower
occupancy and also can appear in multiple conformations accompanied by multiple
configuration of the surrounding water molecules of buffer. Therefore to
localize these low affinity ligands, we need relatively good experimental data
and additional work connected with careful localization of water molecule
networks forming the hydration shell of the protein. It was believed until
recently that protein crystallography cannot describe well the structure of
solvent near the protein surface and thus neither localize well the low
affinity ligands on the protein surface. These difficulties are probably the
reason why crystallographers did not pay attention to these low adhesion
molecules in past.
However, many low affinity ligands have already
been proved as highly efficient tools practically used in health care. Several
tens of studies testing practical usage of these low affinity ligands have been
published every year (drug carriers for safe delivery and release of drugs in
the target tissue, coating materials protecting the biologically degradable
molecules during their transport, control of drug release rate, artificial
additives in food, etc.). In spite of a clear
importance of the subject, our knowledge of how the low affinity ligands
bind to protein surface remains limited until now.
In spite of the fact that polyethyleneglycol
(mostly PEG2000) has been intensively used in crystallization experiments for a
long time [1], only marginal attention
has been paid to soaking of various polymers into the protein crystals and to
crystallographic studies of adhesion between proteins and polymers (low
affinity materials of non-biological origin). Here we show on several examples
[2-10] a number of general problems tackled when one tries to study polymers
and the complex molecular systems mentioned above. Namely we focus on the
problems connected with determination of all conformation states,
conformational freedom of the bound ligand influencing the entropy of the
system, the flipping problem of His, Gln, Asn, determination of water sites in the first hydration shell,
verification of water molecule networks and localization of the relevant parts
of polymer ligands.
It can be said generally that the
experimentally derived view of the molecular structure (the 3D-map of electron
density) contains information about all conformational states and motions of
the molecular system realized during the time of the measurement (usually
several minutes). In the case of a good measurement, parts of the molecular
system remaining stable during the measurement are usually well resolved in the
map of electron density, any single electron can be resolved and the individual
atom positions can be localized with an accuracy up to
The fact that we cannot expect high specificity
of these interactions, because proteins did not pass any genetic selection with
respect to these compounds in past, is on the other hand a great advantage. Molecular
adhesion of such low affinity materials is not critically dependent on a
specific protein, and thus we hope that it will be much easier to generalize
the observations received on a number of experimentally determined structures
of proteins of different origin and to form rules which govern the properties
of these molecular complexes.
The
research is supported by grants of the
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