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SUMMARY:Pipeline for <i>in cellulo</i> structure determination
DTSTART;VALUE=DATE-TIME:20141120T063000Z
DTEND;VALUE=DATE-TIME:20141120T080000Z
DTSTAMP;VALUE=DATE-TIME:20260811T085242Z
UID:indico-contribution-658@events01.synchrotron.org.au
DESCRIPTION:Speakers: Damia Garriga (Monash University)\, Marion Boudes (M
 onash University)\nThe production of diffraction-quality crystals remains 
 the major bottleneck in X-ray crystallography\, as shown by data from the 
 main structural biology consortia. By contrast\, in certain systems\, crys
 tals grow readily in the complex environment of the cell used to express t
 he protein\, be it in the natural context or in a recombinant system for o
 verexpression. Recent interest in these in vivo crystals have \nemerged in
  the context of a growing impact of microcrystallography brought by serial
  microcrystallography at synchrotron and X-ray free electron laser facilit
 ies.\n\nOur aim is to set up a pipeline to streamline in cellulo diffracti
 on\, direct exposure of crystals to X-rays directly through the cells. Usi
 ng in vivo crystals of the recombinant cypovirus polyhedrin\, we show that
  crystal-containing cells could be selectively sorted by flow cytometry ba
 sed on their higher side scattering. Crystal-containing cells were dyed wi
 th Trypan blue to achieve better visualisation\, mounted on micromeshes an
 d flash-frozen. Analysis of these cells on the MX2 beamline of the Austral
 ian Synchrotron show that high-quality diffraction data can be collected f
 rom in cellulo crystals. The structure of the polyhedrin protein determine
 d by molecular replacement closely matches the model previously determined
  from purified microcrystals. Advantages of this approach over conventiona
 l crystallography and future developments will be discussed in the present
 ation.\n\nhttps://events01.synchrotron.org.au/event/3/contributions/658/
LOCATION: NCSS Exhibition Area
URL:https://events01.synchrotron.org.au/event/3/contributions/658/
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