Hung-Jen Wu Research Lab

Chemical Engineering, Texas A&M University

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Research

1. Multivalent lectin binding & bacterial adhesion to host cell membranes

multivalent pathogen_2

Figure 1 Multivalent binding on cell membranes

Glycans expressed on cell surfaces mediate a wide range of biological processes, such as pathogen (e.g. bacterial, viral) and exotoxin adhesion to host cells. Interestingly, a single glycan-protein interaction is often weak; most proteins bind to glycans via multivalent interactions, in which multiple binding domains in a single protein simultaneously interact with multiple glycan molecules. Glycan receptors on cell membranes are often mobile; glycans attached to lipids or membrane proteins can freely diffuse and rotate on 2D fluidic cell membranes, self-organizing to reach multivalent interactions with a protein. Such multivalency leads to higher binding avidity and specificity, and is one of the major recognition principles in many biological systems.

We recently monitored the hetero-multivalent binding process of Cholera Toxin Subunit B (CTB) (i.e. a CTB simultaneously binding to two or more different types of glycolipid receptors). Surprisingly, when a very weak receptor (e.g. GM2) was mixed with the strong binding receptor (e.g. GM1), GM1 activated the interactions between CTB and GM2, enhancing the overall CTB attachment. We demonstrated that the activation of low-affinity receptors was mediated via a simple chemistry mechanism, Reduction of Dimensionality (RD). (see Figure 2) After CTB attaching to the first receptor, the subsequent binding events are confined in a two dimensional membrane surface. Due to the reduced dimensionality of diffusion, the effective concentrations of weak receptors dramatically increase for the subsequent binding events; therefore, a weak receptor can contribute to CTB binding.

The RD mechanism strongly depends on cell membrane dynamics. This presents an issue to conventional ligand-receptor screening assays (e.g. microarray technology, ELISA, etc.) because these tools that often immobilize receptors on solid surfaces miss the inherent membrane dynamics. To date, no theoretical model is available to describe the fundamental RD process. My research group is developing novel sensing technologies and computation tools to investigate the RD mechanism and understand the influence of hetero-multivalency in the pathogenesis of bacteria (e.g. Pseudomonas aeruginosa and Mycobacterium tuberculosis).

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schematics_2Figure 2 Mechanism of Reduction of Dimensionality (RD). A CTB containing five identical subunits binds to two types of glycolipid receptors (GM1 & GM2) on a cell membrane surface. The mechanism entails the following steps: (1) CTB diffuses from solution phase to a membrane surface, and one of the binding sites attaches to one membrane receptor, and (2) Free membrane receptors move two dimensionally, encounter bound CTB, and enable subsequent binding. The reaction rates of subsequent bindings on 2D membrane surfaces are at least 104 times higher than the first binding. Thus, even weak binding receptors can now participate in the second or higher binding events, leading to increased CTB attachment because of the enhancement of effective concentration. This intrinsic mechanism is poorly addressed in most ligand-receptor binding studies.

 

2. Multifunctional nanomaterials for biosensing

My research group focuses on developing highly structured nanomaterials for biosening. We have established the novel instruments to quantitatively explore various biological processes, including nanocube-based lipid bilayer array, multiplexing chemical and biological Raman sensors, and Nanotrap for infectious disease screening.

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nanocube array

Figure 3 Nanocube-based Lipid Bilayer Array. Silica-coated silver nanocubes are covered by supported fluidic lipid bilayers where receptors are presented. Binding kinetics were detected by monitoring absorption spectra with standard microplate reader

3. Quantitative surface enhanced Raman spectroscopy (SERS)

Figure 1

Figure 4. Measuring Reaction Kinetics with Quantitative SERS Using Ag@AuNC Substrate

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4. Role of microvesicle/exosome in infectious diseases

Schematic 2 (for specific aim)

Figure 5. Isolation of Exosome Using Microfluidic Device

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