45 / 101
Biophysics in the Understanding, Diagnosis, and Treatment of Infectious Diseases Speaker Abstracts
40
Post-Translational Modification of a Nucleoid Associated Protein Regulates Cell State in
Mycobacteria
Alex Sakatos, Michael Chase,
Sarah Fortune
.
Harvard TH Chan School of Public Health, Boston, MA, USA.
Although there are many known mechanisms by which eukaryotic cells achieve epigenetic
inheritance, the mechanisms by which prokaryotic cells generate heritable, semi-stable
differences in state remain poorly understood. Here we demonstrate that with a population of
mycobacterial cells, there are privileged subpopulations of cells that are more tolerant of
antibiotic killing than the majority of cells. We show that these differences in cell state are
heritable and semi-stable. We further demonstrate that there is extensive post-translational
modification of the nucleoid associated protein, HupB, at predicted DNA binding residues.
Mutation of the sites of post-translational modification disrupts the formation of subpopulations
of antibiotic tolerant cells. We propose that metabolically driven modification of HupB acts as an
epigenetic regulator, controlling the formation of a specialized subpopulation of antibiotic
tolerant cells.
XChem: From Crystals to Potent Molecules with X-Rays and Poised Synthesis
Frank von Delft
1,2
.
1
University of Oxford, Headington, United Kingdom,
2
Diamond Light Source Research
Foundation, Oxfordshire, United Kingdom.
Fragment-based lead discovery is now a well-established as a powerful approach to early drug or
lead discovery: since small (<250Da) compounds (“fragments”) tend to bind relatively
promiscuously, hits can be readily identified by screening against comparatively small
compound libraries (100s-1000s). What remains challenging is that hits typically bind weakly:
not only must the screening technique be sufficiently sensitive, but potency can only be achieved
through considerable synthetic elaboration. Historically, the most sensitive primary screening
technique of all, direct observation in crystal structures, has been too challenging to be
achievable by but a few labs world-wide. Equally, no consensus has yet emerged on systematic
strategies for synthetic follow-up.
<!--250Da-->Now, beamline I04-1 at Diamond Light Source has established X-ray screening as
a routine medium-throughput experiment with a capacity of up to 500 crystals/day (from soaking
to dataset), a facility being offered to Diamond users since April 2015, with dedicated weekly
beamtime. The highly streamlined process includes image recognition for crystal targeting,
soaking by acoustic dispensing, robot-assisted harvesting, unattended X-ray data collection,
automatic data integration, and pan-dataset electron density analysis for detecting hits. The
technology was developed as a joint research project with the Protein Crystallography group of
the SGC at Oxford University, and has been validated on a series of diverse targets, all of which
have yielded hits.
Moreover, a “poised” fragment library has been developed that provides clear and robust routes
to first-shell follow-up: combined with new algorithms for prioritizing compounds, the ultimate
ambition is to establish how potency can be achieved cheaply from very limited initial
experiments. If achievable, this would have a major impact all aspects of biological research.