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Protein Science Facility

National facility

The Protein Science Facility provides recombinant protein production services using high throughput methods. The facility is focused on His-tagged proteins produced in E. coli and carry out sub-cloning, small-scale expression tests, litre-scale production cultures and a semi-automated two-step protein purification, but also works with tailor made protocols.

In addition PSF provides training and access to instrumentation for macromolecular crystallography and biophysical characterization. Please visit us at ki.se/psf to learn more!

SERVICES

  • Multi construct sub-cloning package. A set of 10 protein variants designed with seven primers, sub cloned into an expression plasmid using LIC cloning.
  • Evaluation and design of constructs
  • Service with ordering of synthetic genes, subcloned into expression vector.
  • Production scale cultures. Usually 1 to 6 (up to 20) litres of TB, induction at 18C over night.
  • Protein purification. Affinity purification utilizing the His-tag followed by a size exclusion chromatography step.
  • Small scale expression and solubility screening. Per cloning package: 1 ml cultures followed by affinity purification and SDS-PAGE analysis.
  • Tailor made additions, e.g. GST affinity, proteolytic affinity-tag removal, extra purification steps such as IEX, additional analyses.
  • Training on biophysical characterisation instruments
  • Training on protein crystallography platform

APPLICATIONS

PSF’s protein production is frequently used by scientists from many Swedish universities as well as companies. Proteins produced by PSF are used for various applications, such as biophysical and biochemical characterization, antibody generation, nanoparticles, pull-downs, crystallization, structure determination, animal experiments, diagnostics, assays and technology development.

The biophysical characterisation instruments are predominantly used for evaluating interactions; proteins with other proteins, DNA/nucleotides, peptides, or small molecule ligands. Buffer screens and ligand screens are often performed in order to find stabilizing conditions and affinity constants are often sought in combination with structure determination of complexes.

The macromolecular crystallography set-up offers use of robotics for liquid handling and crystallisation, imaging systems and microscopes for crystal visualisation, tools for crystal handling and mounting. There are also computer resources with crystallographic software, super computer access, and remote synchrotron data collection.

EQUIPMENT

  • Standard protein production instrumentation.
  • Isothermal titration calorimetry. ITC200
  • Thermal melt assay using differential fluorescence spectroscopy. DFS
  • Thermal melt assay using StarGazer.
  • Circular dichroism. CD
  • SPR. Biacore 2000
  • Protein crystallography set-up
  • Liquid handling robots, TECAN, Mosquito, Phoenix
  • Imaging systems, Formulatrix
  • Crystal handling and freezing

RECENT PROJECTS

Please see our list of publications related to PSF