VIB Protein Service Facility, UGent
Expert Technologist : Jurgen Haustraete
Current team members:
Chantal Eichperger, Jannick Leoen, Jurgen Haustraete, Maarten Hofkens
The study of proteins
If DNA can be considered life’s software, proteins are the hardware. Characterizing proteins and their behavior often needs the availability of sufficient amounts of purified protein, which can be used for biochemical studies, characterization and measurement of interactions, use as an antigen to obtain antibodies, structural determinations, or use as an experimental therapeutic.
The VIB Department for Molecular Biomedical Research (DMBR, UGent) has a long history on developing and using recombinant expression technology, resulting in an established track record with both internal DMBR-projects and projects for industrial partners. This expertise has been formalized in a structured core facility combining services for expression (upstream), production optimization and protein purification (downstream).
Every step in protein expression and purification
The PSF allows 43 researchers to outsource either every single step in the protein expression and purification process, or a complete project starting from the cDNA sequence to purified protein. The Protein Service Facility (PSF) has a capacity to grow microbial cells (E. coli and P. pastoris) to 20 L and mammalian cells (mainly transient HEK293T production) to 5 L scale. Equipment for downstream processing (lysis, collection, UF/DF, chromatography) is in place to accommodate this production scale. This scale allows production in the 1 mg – 1 gram range, depending on the protein.
Starting with a cDNA sequence, or a cDNA sequence that expresses protein problematically, the PSF offers a number of services to come to a convenient solution:
• FastScreen: Parallel cloning and expression in a subset of different expression systems for E. coli. This system is optimized for (superior) promoter regulation, mRNA stability, transcription termination and plasmid stability. Variations are possible in order to optimize the translation initiation, the codon usage, the signal sequence, the addition of detection tags and fusion partners. To keep the protein soluble and active, it is sometimes necessary to express the protein of interest linked to a fusion partner. Numerous fusion partners (cytoplasmic and secreted) are assembled in the system for side by side comparison. Three different protease systems can be chosen in the system to optimize the cleavage of your fusion protein. A FastScreen service offers an alternative to iterative approaches, all in a standardized vector system for a non-biased comparison. The E. coli screening can be expanded by evaluating other hosts such as Pichia pastoris (also with native and fusion protein approaches), or production in mammalian cells. In vitro transcription / translation systems can also be used for optimization of translation initiation, or for rapid evaluation of activity of protein mutants. The whole Fastscreen procedure screen usually takes 4 weeks.
• Fastfold: A matrix approach to optimizing refolding conditions and recovery of proteins from inclusion bodies. A typical FastFold optimization will take about two weeks to perform (analyzing both refolding conditions and refolding method).
• Fastclean: If no purification method is available, a screen can be set up to test a range of different (or combinations of) matrices to purify the protein of interest. A typical FastClean optimization will take about 2 weeks to perform.
The PSF
The PSF is operated by 4 technicians specialized in the necessary technologies. The projects and the facility are managed by Jurgen Haustraete. The PSF offers transparent project planning and follow up of each service.
Interested?
Contact Jurgen Haustraete at the PSF
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