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BIACe: Belgian Instruct Affiliate Centre

Instruct-BE

Instruct-BE

Protein production: The BIACe protein production and purification platform encompasses genetic engineering, high throughput mutagenesis and cloning in a variety organisms and cell lines (bacterial strains, yeast, filamentous fungi, SF9, CHO), analytical or pilot scale recombinant protein production (shaking incubators and 1L to 60L fermentors for batch and fed-batch cultures), cell harvesting or supernatant cleaning using continuous centrifugation or hollow fiber filtration, cell disintegration to recover proteins produced in the intracellular compartment and protein purification at the analytical and pilot scales. Furthermore, we are currently developing a high throughput mutagenesis platform, based on a Microlab Star Hamilton automated liquid handling workstation.

NMR, X-ray crystallography and solution scattering: BIACe provides access to three dedicated liquid handling systems for crystallization screen formulation and plate and nanodrop dispensing, a visualization system. X-ray technologies encompassa state of the art microfocus rotating anode generator equiped with single crystal X-ray diffraction set up on one port and SAXS set up on a second port. The consortium has regular access to the European synchrotron facilities. Through the Jean Jeener NMR centre, the consortium offers access to 600 and 800MHz magnets for biomolecular NMR.

Biophysics: BIACe offers a large range of equipment and expertise for protein structural characterization: ITC, DSC, CD, fluorescence spectroscopy, atomic force spectroscopy, microscale  thermophoresis, SRP, stopped flow & quenched flow and infrared spectroscopy (see below).

Unique structural biology platforms offered to the International Instruct community:

Nanobody-assisted structural biology: Nanobodies are antibody-derived single domain proteins that contain the unique structural and functional properties of naturally-occurring heavy chain only antibodies. By rigidifying flexible regions and obscuring aggregative surfaces, nanobody complexes warrant conformationally uniform samples that are key to protein structure determination by X-ray crystallography. The elucidation of the first GPCR structure in its active state using a conformationally selective Nanobody demonstrates the power of the Nanobody platform to generate diffracting quality crystals of the most challenging targets including membrane proteins, flexible proteins, aggregating proteins and protein-protein complexes.

BIACe will offer Instruct users access to its platform to generate Nanobodies that facilitate the structural investigation of proteins that are recalcitrant to purification and crystallization. Future innovative applications of Xaperones in in situ fluorescence imaging, mass spectrometry, NMR, X-ray crystallography, electron microscopy and tomography, X-ray imaging or other relevant biophysical methods will be explored in an integrated fashion within the most multidisciplinairy projects selected by the Instruct framework.

Fourier Transform Infrared Spectroscopy: FTIR provides an extremely precise fingerprint of protein structure. While this is not per se a high resolution method, it has become a method of choice for the study of protein structure dynamics in complex environments. We developed methods, applications and softwares in this specific field. FTIR allows to monitor protein structural parameters (secondary structure, orientation of secondary structures in membranes and H/D exchange kinetics) while the environment is modulated, both in defined systems or in whole cells. Because the FTIR spectrum reflects the folding state of a protein and distinguishes easily between different folds even with identical secondary structure content, this method can also be used for protein quality control and the study of protein dynamics. Finally, the IR microscope can examine the smallest crystals and identify its nature (protein or other). 

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