Chemistry Reference
In-Depth Information
Figure 4.3
STD competition experiments using a compound with known binding site to rank
the binding affinity of fragment hits or fragment analogues to a target protein. (A) and (B) are
the aromatic parts of STD spectra of two different fragment hits in the absence of competing
compound. In the STD spectra (C) and (D), the competing compound is present in equimolar
amounts compared to the fragment hits, and in (E) and (F), the competing compound is present
in excess over the fragment hits. From this series of experiments, it is possible to conclude that
the fragment hit in the STD spectra to the left (A, C and E) binds to the target with a higher affinity
than the fragment hit in the STD spectra to the right. Further, since it is possible to displace
fully both fragment hits with the competing compound, the fragment hits bind specifically to
the target protein and, most probably, at a site overlapping with the competing compound.
4.6 Cases
This section provides descriptions of two cases of fragment-based screening campaigns
performed at Biovitrum. The first case, where the adipocyte fatty acid binding protein
(A-FABP) was the target protein, is described in some detail,
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whereas the second example
is described in more general terms. Both target proteins (A-FABP and the Ser/Thr kinase)
were subjected to both HTS and fragment screening by NMR. The fragment screening
started at the same time as assay development for HTS was initiated, i.e. as soon as soluble
target protein had been produced. Since no assay development or formatting is necessary to
start fragment screening by NMR, the results presented here were available before the HTS
was completed. The goal with the fragment screenings was to find soluble compounds with
low micromolar or better potencies and high ligand efficiencies. The further development
of these compounds was then performed together with the HTS hits by the respective
project teams.
