Glycan recognition
Interesting aspect of glycan-mediated interactions is the recognition of glycan clusters as a specific ligand for protein binding. Such clustered saccharide patches (CSPs) may form on heavily glycosylated proteins (A), on cell membranes because of interactions between two or more glycosphingolipids (B) or glycoproteins (C). CSPs can also form on pathogen polysaccharides (D). Glycosynapses are clustered glycan microdomains, which mediate cell–cell and cell–extracellular matrix adhesion and signaling (E). Arrows indicate likely CSPs.
All cells in nature are covered with a dense and complex array of glycan (sugar) chains. Specific recognition and binding of glycans is critical aspect of cellular interactions, both within and between species. Glycan-protein interactions tend to be of low affinity but high specificity, typically utilizing multivalency to generate the affinity required for biologically relevant binding.
M. Cohen and A. Varki.
Int Rev Cell Mol Biol, (2014)
In addition, cells can be covered with a layer of pericellular coat that comprise of hyaluronan. Hyaluronan is a unique glycosaminoglycan polymer that is not sulfated and is not attached to a protein. Hyaluronan binds receptors on the cell membrane and extends to form micrometer-thick halo around the cells. The pericellular coat mediates the initial step of cell adhesion, and in oocytes (eggs) it forms a barrier that sperm has to penetrate in order to fertilize the egg. It is also an integral component of the extra cellular matrix microenvironment.
M. Cohen et al. ChemBioChem, (2004)
Influenza infection and mucins
Influenza viruses are significant human respiratory pathogens that cause seasonal infections and occasionally cause pandemics. It is an enveloped virus belongs to the family of Orthomyxoviridae. Influenza subtype is classified based on the two major envelope viral proteins, one that binds to sialic acids (hemagglutinin or H) and the other that cleaves sialic acids (neuraminidase or N). These viruses infect their hosts by binding to sialoglycans on airway (mammals) or gut (birds) epithelia. Hemagglutinin facilitates infection by binding to the epithelia cells, and the enzymatic activity of viral neuraminidase is required to enable escape of progeny virions from host cells. However, both target tissues are covered with a thick layer of mucus that presents abundance of highly sialylated mucin glycoproteins. Viral hemagglutinin binds to the sialylated mucins resulting virus entrapment in the mucus layer, and neuraminidase activity enables release of trapped virus form the mucus layer.
We have demonstrated that neuraminidase inhibitors, such as oseltamivir (Tamiflu) enhance the ability of mucus layer to protect underlying cells from infection. We are currently developing a platform to detect potential neuraminidase inhibitors that enhance the natural ability of the mucus layer to protect against influenza infection (patent pending).
3D mucin biomimetics array
Virus binding specificity is determined by the sialic acid type and by its linkage to the underlying glycan structure. For example, human influenza viruses preferably bind alpha-2-6 sialoglycans, which are abundant in the upper airways of humans. In contrast, avian influenza viruses prefer sialic acids in alpha-2-3 linkage, found in the birds gut and in human lower respiratory tract. Therefore changes in hemagglutinin binding specificity may indicate newly gained virus ability to infect a new host. This is a major concern in particular changes in the binding specificity of highly pathogenic avian influenza viruses.
We developed a mucin-biomimetic beads array for high-throughput screening of influenza virus binding specificity. This array is comprised of a magnetic bead core coated with synthetic mucin-like polymers that present a dense array of glycan structures. Similar to other glycan-binding proteins, multivalent interactions are required to facilitate virus binding to the host. Our array presents glycans at a density that is comparable to hemagglutinin density on the virus envelope. In collaboration with the Kamil Godula and from UC San Diego, and Walter Boyce from UC Davis, we are monitoring binding specificity of whole influenza virus in an effort for early detection of strains that have a potential to cause pandemics.
The mucin-biomimetic beads array is a useful tool, and can be applied for testing glycan-binding specificity of other glycan-binding viruses and bacteria. Furthermore, the mucin-biomimetic beads can be used to isolate virus and bacteria from biological samples. I am currently exploring various applications for the array including microbiome studies, food pathogens and different viruses.
