Deciphering the structural attributes of protein–heparan sulfate interactions using chemo-enzymatic approaches and NMR spectroscopy

Heparan sulfates (HS) is a polysaccharide found at the cell surface, where it mediates interactions with hundreds of proteins and regulates major pathophysiological processes. HS is highly heterogeneous and structurally complex and examples that define their structure–activity relationships remain limited. Here, in order to characterize a protein–HS interface and define the corresponding saccharide-binding domain, we present a chemo-enzymatic approach that generates 13C-labeled HS-based oligosaccharide structures. Nuclear magnetic resonance (NMR) spectroscopy, which efficiently discriminates between important or redundant chemical groups in the oligosaccharides, is employed to characterize these molecules alone and in interaction with proteins. Using chemokines as model system, docking based on NMR data on both proteins and oligosaccharides enable the identification of the structural determinant involved in the complex. This study shows that both the position of the sulfo groups along the chain and their mode of presentation, rather than their overall number, are key determinant and further points out the usefulness of these 13C-labeled oligosaccharides in obtaining detailed structural information on HS–protein complexes.

Lectins-Robust and Quintessential Proteins of Nature

Recent advances in glycobiology have proclaimed that cell surface saccharides play a pivotal role in recognition events. More accurately, these saccharides may be complexed by lectins. Lectins are saccharide binding proteins that are highly specific for sugar moieties of other molecules. They are ubiquitous in nature. They are diverse proteins with spectacular potential and overwhelming medical applications. Hence, there is quest for identification of new lectins and their precise mechanism of action with specific ligands. In this review, main features of lectins are succinctly described with emphasis on ligand binding and therapeutic applications.

Major conformational changes in the structure of lysozyme obtained from a crystal with a very low solvent content

A new crystal form of lysozyme with a very low solvent content (26.35%) has been obtained in the orthorhombic space group P212121 (with unit-cell parameters a = 30.04, b = 51.68, c = 61.53 Å). The lysozyme structure obtained from these crystals does not show the typical overall fold. Instead, major conformational changes take place in some elements of the secondary structure and in the hydrophobic core of the protein. At the end of the central α-helix (α2), Glu35 is usually buried in the catalytic site and shows an abnormally high pK a value, which is key to the activity of the enzyme. The high pK a value of this glutamate residue is favoured by the hydrophobic environment, particularly by its neighbour Trp108, which is important for structural stability and saccharide binding. In this new structure, Trp108 shows a 90° rotation of its side chain, which results in the rearrangement of the hydrophobic core. Conformational changes also result in the exposure of Glu35 to the solvent, which impairs the catalytic site by increasing the distance between Glu35 and Asp52 and lowering the pK a value of the glutamate. Altogether, this new lysozyme structure reveals major conformational changes in the hydrophobic core and catalytic site that might play a role in the folding and bactericidal function of the protein.

Structures of the transcriptional regulator BgaR, a lactose sensor

The structure of BgaR, a transcriptional regulator of the lactose operon inClostridium perfringens, has been solved by SAD phasing using a mercury derivative. BgaR is an exquisite sensor of lactose, with a binding affinity in the low-micromolar range. This sensor and regulator has been captured bound to lactose and to lactulose as well as in a nominal apo form, and was compared with AraC, another saccharide-binding transcriptional regulator. It is shown that the saccharides bind in the N-terminal region of a jelly-roll fold, but that part of the saccharide is exposed to bulk solvent. This differs from the classical AraC saccharide-binding site, which is mostly sequestered from the bulk solvent. The structures of BgaR bound to lactose and to lactulose highlight how specific and nonspecific interactions lead to a higher binding affinity of BgaR for lactose compared with lactulose. Moreover, solving multiple structures of BgaR in different space groups, both bound to saccharides and unbound, verified that the dimer interface along a C-terminal helix is similar to the dimer interface observed in AraC.

Charge reversal and swelling in saccharide binding polyzwitterionic phenylboronic acid-modified poly(4-vinylpyridine) nanoparticles

We present the synthesis and characterization of zwitterionic poly(4-vinylpyridine) nanoparticles quaternized with phenylboronic acid (QxPVP-PBA) whose size and surface charge can be tuned by varying the saccharide and the degree of quaternization.