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SUMMARY:Contr. talk - Towards Neutron crystallography of membrane proteins
 : Insights into production of deuterium-labelled OmpF
DTSTART;VALUE=DATE-TIME:20210526T074000Z
DTEND;VALUE=DATE-TIME:20210526T080000Z
DTSTAMP;VALUE=DATE-TIME:20260722T000810Z
UID:indico-contribution-1137@lindico453.srv.lu.se
DESCRIPTION:Speakers: Swati Aggarwal (European Spallation Source)\nHydroge
 n bonds play a crucial role for protein function and involved in almost ev
 ery mechanism from foundation of protein structure to enzyme catalysis. Hy
 drogen (1H) atoms form the basis of hydrogen bond that is not scattered by
  X-ray crystallography due to its poor scattering power. Neutron protein c
 rystallography (NPX) is a powerful tool that is capable of locating hydrog
 ens and study the significance of hydrogen bonding interactions in biomacr
 omolecules. However\, due to\nthe requirement of large crystals very few n
 eutron structures have been deposited in PDB with no membrane protein stru
 cture determined yet. Additionally\, 1H has a negative scattering length a
 nd large incoherent cross-section giving rise to a significant background 
 noise in neutron data collection.\nThis effect can be minimized by isotopi
 c substitution of 1H with its heavier isotope deuterium (2H or D) leading 
 to less ambiguous data analysis and better structure interpretation. Overa
 ll ˜25% H atoms in a protein are solvent exchangeable and can be exchange
 d by dissolving in heavy water. However\, complete deuterium labelling (pe
 rdeuteration) is required for the remaining 75% H atoms. In this work\, an
  optimized methodology for large scale production of perdeuterated bacteri
 al outer membrane protein F (OmpF) has been designed. OmpF was produced in
  deuterated minimal medium with different carbon sources. Mass spectrometr
 y and thermal stability experiments\nverified the purity and level of deut
 eration of OmpF protein. Perdeuterated OmpF crystals also diffracted X-ray
 s to 9 Å resolution emphasising the need of fine tuning of perdeuterated 
 crystallisation conditions.\n\nhttps://lindico453.srv.lu.se/event/219/cont
 ributions/1137/
LOCATION:
URL:https://lindico453.srv.lu.se/event/219/contributions/1137/
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