Funding:

© CIC biomaGUNE 2013

» Credits

HomeResearch Biosurfaces

People

back

Research Units

Biosurfaces

Laboratory 3

The self-organization of molecules into dynamic and hierarchical supramolecular assemblies is a key feature of biological structures. The resulting architectures exhibit new qualities that are distinct from those that characterize its individual components. Our group is particularly interested in two types of assemblies: lipid membranes and the gel-like, polysaccharide-rich coats as they surround many cells. For a thorough investigation of the physical principles underlying the structure and function of these architectures, it is desirable to move from living cells with their complex dynamics to well-controlled models with tunable complexity. We create such model systems on solid supports. Modern techniques of surface nanostructuration and biofunctionalization are employed to guide the assembly down to the nanometer-scale. For the characterization of such model systems, we develop and use a toolbox of biophysical in situ characterization techniques, including quartz crystal microbalance with dissipation monitoring (QCM-D), atomic force microscopy (AFM), reflection interference contrast microscopy (RICM), ellipsometry and fluorescence methods. The created structures are employed as specialized platforms for biosensors and for the control of cellular fate.

Person in charge

Members

Projects

Biomolecular Hydrogels - From Supramolecular Organization and Dynamics to Biological Function

This Project is funded by an ERC Starting Grant .

Weblink to project

Model Systems of the Pericellular Coat

Many cells equip themselves with a gel-like coat that is rich in the polysaccharide hyaluronan. This pericellular coat can reach a thickness of several micrometers. It is invisible in common light microscopy, since its extreme hydration renders the optical contrast very low. The grafting of hyaluronan to the cell membrane and its interaction with hyaluronan-binding molecules can give rise to different supramolecular structures, such as cross-linked networks or polyelectrolyte brushes.

 

By reconstituting these intriguing structures in vitro, the physico-chemical properties of such coats can be quantified. We create model systems of the pericellular coat that are based on supported lipid membranes. This immobilization platform allows us to create hyaluronan layers with well-defined composition and structure, such as brushes of end-grafted hyaluronan.

Under inflammatory conditions and ovulation, hyaluronan-rich coats undergo significant remodeling. The inflammation-associated protein TSG-6 was hypothesized to be implicated in the coat remodeling by cross-linking HA chains. With the aid or our model systems, we could provide evidence that TSG-6 can indeed act as an effective HA cross-linker, and shed insight into the cross-linking mechanism. The cross-linking units are TSG-6 oligomers which form upon binding of TSG-6 to HA. TSG-6 incorporation at physiologically relevant concentration collapses and rigidifies HA films. Cross-linking might hence influence the mechanical environment of cells and the local remodeling of the glycocalyx might also serve as a signal for leukocyte recruitment to sites of inflammation.

Interaction of Hyaluronan with the Cell Surface

CD44 is a major cell surface receptor for hyaluronan (HA). It is found on many cell types, and of particular importance in inflammation-like processes and tumor metastasis. The interaction of hyaluronan with the cell surface is thought to be stabilized by multivalent interactions between polymeric HA and several cell surface receptors. The molecular mechanisms behind the regulation of binding, and in particular the different biological functions that are elicited by HA of different molecular weight, are currently not well understood.

 

We have designed tunable in-vitro model systems that mimic the cell surface in the sense that the HA binding domain of CD44 is immobilized in its native orientation and at controlled density to a supported lipid bilayer (SLB). Employing techniques such as QCM-D, ellipsometry and microinterferometry and concepts from polymer theory, we analyze qualitatively and quantitatively, how the multivalent presentation of CD44 on the membrane surface regulates the binding and organization of hyaluronan and its complexes.

The Permeability Barrier of Nuclear Pore Complexes

All macromolecular transport between the cytosol and the nucleosol of living cells occurs through nuclear pore complexes (NPCs). The transport is selective: objects beyond a certain size (30kDa) need to attach to soluble nuclear transport receptors (NTRs) in order to be channeled efficiently through the pore. A supramolecular assembly of specialized and natively unfolded protein domains  (FG repeat domains) within the NPC is thought to be the key component of the NPC´s permeability barrier. The mechanism behind transport selectivity is at present only poorly understood.

 

We design tailor-made nanoscale model systems of the permeability barrier of NPCs, such as thin films of FG-repeat domains that are end-grafted to planar solid supports. The physico-chemical properties of the films and their selective interaction with NTRs and other probes are characterized in detail by a combination of biophysical surface-sensitive techniques. We employ the nanoscale model systems to gain insight into the inter-relationship between supramolecular structure and function of the permeability barrier.

Nanoparticles & Acousto-Mechanical Sensing

Quartz crystal microbalance with dissipation monitoring (QCM-D) has become a popular tool to investigate biomolecular adsorption phenomena at surfaces. In contrast to optical mass-sensitive techniques, which commonly detect the adsorbed nonhydrated mass, the mechanically coupled mass measured by QCM-D includes a significant amount of water. A mechanistic and quantitative picture of how the surrounding liquid couples to the deposited molecules has long been elusive for apparently simple phenomena like the random adsorption of proteins or other nanometre-sized particles on a planar surface.

 

We employ in situ combinations of QCM-D with other sensing techniques (in particular ellipsometry) and theoretical modelling to elucidate this question. The insights are used for the development of novel sensing applications. With this methodology, it is for example possible to detect the clustering of proteins on supported lipid bilayers without any labels. 

Dynamic Tuning of Model Membrane Lipid Composition

We have developed an efficient and robust method to dynamically modulate the glycolipid content in supported lipid bilayers. The method is based on glycolipid transfer protein, a soluble protein that can selectively transfer glycolipids from one membrane compartment to another. It provides quantitative control on the lipid composition of the bulk facing bilayer leaflet. Owing to the importance of glycolipids in the function of cell membranes and membrane proteins, the method may find widespread use in membrane research.

Publications

publications list

Additional information