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Metals in Medicine Network NSW Regional Meeting

Metals in Medicine Network NSW Regional Meeting. Structural Biology Mitchell Guss School of Molecular & Microbial Biosciences University of Sydney. 19 Jan 2004. Structural Biology in Australia. Limited to X-ray and NMR of macromolecules (may not be accurate) NSW (X-ray:2*, NMR:3(4))

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Metals in Medicine Network NSW Regional Meeting

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  1. Metals in Medicine Network NSW Regional Meeting Structural Biology Mitchell Guss School of Molecular & Microbial Biosciences University of Sydney 19 Jan 2004

  2. Structural Biology in Australia • Limited to X-ray and NMR of macromolecules (may not be accurate) • NSW (X-ray:2*, NMR:3(4)) • ACT (X-ray:1, NMR:1) • VIC (X-ray:5, NMR:2) • QLD (X-ray:3(4), NMR:2) • SA (X-ray:1, NMR:1) • WA (X-ray:1, NMR:1) *sites not PI’s

  3. Sydney: Historical Outline • 1954-1982: Metal amino-acid and metal peptide complexes (Chemistry: Freeman) • 1972-1993: Metalloprotein structures (Chemistry: Freeman, Guss & Wright) • 1994-current: Metalloprotein structures (MMB: Freeman, Guss, Mackay, Maher, Matthews)

  4. Selected Highlights • Blue copper proteins • Plastocyanin (1st new protein structure solved in Australia) • Cucumber Basic Protein (1st metalloprotein structure solved by the multiple wavelength anomalous scattering method) • Auracyanin • Iron proteins • Legume hemoglobin • Rubredoxin (mutants and different metals) • Purple acid phosphatase • Copper enzymes • Amine oxidases (pea seedling, Arthrobacter globiformis, Pichia pastoris) • Others • Aminopeptidase P (manganese) • Prolidase (cobalt, zinc) • Selenate reductase (selenium) • Dihydroorotase (zinc) • Transcription Factors (zinc) • Oligonucleotides (cobalt)

  5. Principal Techniques • Nuclear magnetic resonance spectroscopy (Mackay, Matthews, Kuchel) • 400 MHz, 600 MHz (MMB, cryoprobe 2004/5) • 800 MHz (ANU, 2004) • X-ray crystallography(Collyer, Freeman, Guss, Maher) • Rotating anode generator, Marresearch image plate detectors (MMB) • Synchrotron (SSRL, APS)

  6. Facilities for Structure Characterisation - MMB • Ultracentrifugation • Plasmon resonance • CD spectroscopy • Microcalorimetry • Mass spectrometry (Electrospray and Maldi) • Fluorescence spectroscopy • Dynamic and multi-angle light scattering

  7. Additional Facilities - MMB • Protein expression and purification (Fermentation for bacterial and mammalian* cells) • Capillary electrophoresis • FPLC and HPLC • Proteomics (2D gels, automated spot picking, mass spectrometry) • Animal house • PC2 & PC3 labs *only bacterial expression to date

  8. Computing - MMB • X-ray & NMR groups share a mixed LINUX/IRIX cluster • 6 Intel-based LINUX servers providing file and compute resources (100 GBytes user and 200 GBytes of scratch storage; 10 cpus) • 6 LINUX workstations; 7 IRIX (SGI-O2) workstations for graphics applications

  9. Collaboration or Service? • Protein structure determination is becoming increasing “routine”: • Still requires major commitment of personnel resources • Not a service • Close collaboration • Growing focus on the biology of systems • Use of whatever techniques required to solve the problem • Lack of technical support staff to enable routine (casual) use of instrumentation

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