Difference: TemimpsReports (12 vs. 13)

PSI Annual Meeting: Dec 8-9, 2010

TEMIMPS Goals for 2011

• Technology development will dominate 2011 as we establish the tools necessary for attacking our protein targets
  • Establish pipeline for protein production from bacteria, yeast and insect cells
  • Establish standard conditions for screening 2D crystallization of novel targets
  • Implement methods for detergent removal/crystallization
    • microfluidic 96-well dialysis block
    • robotic micropipettor for incremental addition of cyclodextrins
  • Develop software for automated imaging and evaluation of crystallization trials by electron microscopy
  • Demonstrate utility of phase extension methods for electron crystallography
  • Implement iterative helical reconstruction methods

• Targets
  • P2A3 Zn transporter: Comparison of membrane bound form with X-ray structure of detergent-solubilized homolog (YiiP^?)

TEMIMPS Goals for next 5 years

• Targets
  • Existing crystals
    • AQP2: Comparison of constructs mimicking phosphorylated/dephosphorylated state
    • AQP0: Interaction of channel with native lipids
    • KvAP^? channel: Structure of membrane bound channel to address gating
  • Novel targets
    • GPCRs: cannabinoid, seratonin
    • TRP channnels: TRPA1 pain receptor
    • Rhomboids
    • secondary transporters: Heme transporter FLVCR
  • Oligomeric proteins
    • gamma-secretase
    • Kdp (ATP-dependent P-type potassium pump)
  • Complexes
    • G-protein in complex with GIRK channel
    • AQP2 in complex with AKAP79 adaptor
    • Two-component system (PhoP^?/PhoQ)

• Technologies
  • establish general high-throughput pipeline for screening 2D crystallization within lipid bilayers
    • automation of crystallization
    • automation of imaging
    • integration with database
    • general enough and affordable to interest other EM groups
  • develop software for automated data collection
    • low-dose imaging
    • electron diffraction
  • improve software for structure determination
    • novel methods for image analysis of poorly ordered crystals
    • more robust/automated analysis of electron diffraction

• Protocols
  • define important parameters affecting 2D crystallization
    • define matrix of conditions for crystallization screen
  • extend methods for structure determination by electron crystallography
    • molecular replacement
    • phase extension
    • density modification

Notable achievements

• none to date

• Set ALLOWTOPICVIEW =

-- DavidStokes - 14 Nov 2010