12
X-ray Crystallography
Considered the most sensitive screening technique in
FBDD, it's also the only method that can provide details of
the ligand binding site and the mode of binding. It's also the
least susceptible to false positives.
X-ray crystallography requires growing crystals of the target protein
that are soaked in fragment samples at concentrations of up to 100
mM. Crystals are then rotated in a beam of X-rays to generate an
electron density map capable of producing a three-dimensional model
of the protein and any bound fragment. Formerly, X-ray crystallography
was used to determine structure only as part of the optimization
process a er hit detection was performed by other methods. But
currently, it can be used as a high-throughput, front-line method for
screening hits. Producing workable protein crystals, however, can
be challenging and crystallized proteins may not be able to assume
the same conformations in solution. Thus, there is a potential for
obscuring binding sites resulting in false negatives.
Most sensitive and best for
providing details of binding
site and mode of binding
Crystallization of proteins
remains challenging
DWhen
affinities
become tight,
one must also
be very careful
to optimize the
regeneration
conditions of
the chip to
