The Brussels based Nanobodies4Instruct center produces and characterizes Nanobodies to be used as auxiliary tools in structural and cellular biology
Nanobodies are the small (15 kDa) and stable single-domain fragments harboring the full antigen-binding capacity of camelid heavy chain–only antibodies. Nanobodies are exquisite chaperones for crystallizing membrane proteins, multiprotein assemblies, transient conformational states and intrinsically disordered proteins. Nanobodies can also be used for other applications in structural biology. Domain-specific Nanobodies have been used in single-particle electron microscopy (EM) to track these domains in particle projections. Because Nanobodies can be functionally expressed as intrabodies in eukaryotic cells, these single-domain antibodies can also be used to track their targets inside a living cell.
The Nanobodies4Instruct centre generates conformational Nanobodies to facilitate the structural analysis of proteins that are notoriously difficult to purify, to crystallize or to study by another method. Collective efforts of several laboratories have demonstrated that Nanobodies are exquisite chaperones for crystallizing complex biological systems such as membrane proteins, transient multiprotein assemblies, transient conformational states and intrinsically disordered proteins. Further, they can be used as structural probes of protein misfolding and fibril formation.
Nanobodies are the small (15 kDa) and stable single-domain fragments harboring the full antigen-binding capacity of the original heavy chain–only antibodies that naturally occur in camelids. Nanobodies are encoded by single gene fragments, they are easily produced in microorganisms and they exhibit a superior stability compared with derivatives of conventional antibodies such as Fabs or scFvs. Because of their compact prolate shape, Nanobodies expose a convex paratope and have access to cavities or clefts on the surface of proteins.
Innovative applications of Nanobodies in in situ fluorescence imaging, mass spectrometry, NMR, electron microscopy and tomography, X-ray imaging or other relevant biophysical methods will be explored in an integrated fashion within the most multidisciplinary projects selected by the Instruct framework. Because many Nanobodies can be functionally expressed as intrabodies in eukaryotic cells, these single-domain antibodies can also be used as biosensors to track conformational properties of their targets inside a living cell.
The Nanobodies4Instruct centre is an initiative of the Steyaertlab. The Steyaertlab is located in Brussels and is part of the Structural Biology Research Center (SBRC) at the Vrije Universeit Brussel (VUB) and the Vlaams Instituut Biotechnologie (VIB). Nanobodies4Instruct is supported for the Flemish Participation in ESFRI by the Research Foundation - Flanders (FWO),
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.
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).
Our labs our on the grounds of the Etterbeek Campus of the Vrije Universiteit Brussel (VUB). We are located close to downtown Brussels and can be easily reached by train, tram, metro and car. All our labs and offices are in the E-building, on the 7th floor. In case you've never been here, the maps below will help you out.
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