Native state electrospray
ionization mass spectrometry (ESI-MS) is, in theory, a fast and easy way to
find fragments: just mix protein with fragment, shoot it on the MS, and look
for complex. As a bonus, the exact mass tells you whether your fragment is what
you think it is (or at least whether it has the right mass). However, published
examples are relatively rare, and not always favorable. A new paper in J. Med. Chem. by Tom Peat, Sally-Ann
Poulsen, and their colleagues at CSIRO and Griffith University seeks to change
this.
The researchers chose the fragment-friendly
model protein carbonic anhydrase II (CA II) as their target. They first
screened a library of 720 fragments, each at 100 µM, using surface plasmon
resonance (SPR). This yielded 7 hits, with affinities ranging from 1.35 to 1280
µM. These seven hits were then assessed by ESI-MS using equimolar
concentrations of protein and fragment (10 or 25 µM each). Encouragingly, all
seven hits confirmed. Soaking these fragments into crystals of CA II yielded
structures for six of them.
This is nice, but of course the
real question is how well ESI-MS works as a primary screen. To address this,
the researchers chose 70 compounds structurally related to the 7 hits and
independently tested these using both SPR and ESI-MS. This yielded 37 hits, of
which 24 were detected both by SPR and ESI-MS. In fact, every SPR hit was confirmed by ESI-MS. Of 14 fragments subsequently
soaked into crystals of CA II, 7 provided interpretable electron density.
This is impressive, and the
researchers note that the level of agreement between SPR and ESI-MS might be
better still, since some of the ESI-MS hits did give signals by SPR – they were
just weaker than the chosen cutoff (KD ≤ 3 mM). Thus, in contrast to
a paper discussed last year, ESI-MS does seem to be a sensitive detection
method. In fact, given the low concentration of fragment needed, the
researchers suggest that it could be useful for screening fragments with lower
solubilities.
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