**2. Multivalent interaction**

The saccharide-protein interaction plays important roles in the living system, and the novel biomaterial fabriction is expected using the interaction. However, the saccharideprotein interaction is basically weak, and it is difficult to utilize and detect the interac‐ tions. It has been reported that the saccharide-protein interaction can be amplified by the multivalency [6, 7, 8]. Actually, saccharides on the cell-surfaces are displayed in a multi‐ valent manner. The glycolipids form densely saccharide structures of lipid-rafts [9], and glycoproteins usually have multivalent saccharide structures, which provies the multiva‐ lent saccharide-protein interactions.

**Figure 1.** Schematic illustration of multivalent saccharide compounds.

The artificial multivalent saccharide displays also enables the multivalent interaction be‐ tween saccharide and protein. Various artificial compounds with multivalent saccharides have been reported (Figure 1). Proteis are commonly used as carries for the multivalent pre‐ sentation of antigens, and bovine serum albumin (BSA) is the representative [10]. Peptides are used as a scaffold of saccharide display [11]. Saccharide conjugates with DNA [12], cy‐ clodextrin [13] and polymers have been also reported to exhibit multivalent interactions. Saccharide conjugtes with peptides and proteins are appropriate structure for phamaceuti‐ cal substances because of the biocompatibility and the fine structures. The glycopeptides to‐ ward shiga toxins (toxins from *E.coli* O-157 and enterohemorrhagic *E.coli*), influenza virus [14] and lectins [15] were reported.

### **2.1. Glycopolymer**

The saccharide-protein interactions are also important in terms of protein analyses (pro‐ teome), because the interaction is important to clarify the biological function of proteins. [5] The saccharide immobilized substrates are investigated for the saccharide-microarray. In ad‐ dition, the saccharide-protein interactions is a potential markar of various diseases like in‐ fection of pathogens (e.g. viruses, bacteria, Cholera, and Shiga toxin) and cancer. Therefore,

An Integrated View of the Molecular Recognition and Toxinology - From Analytical Procedures to Biomedical

In this chapter, we describe the materials with molecular recognition ability of sugars. Section 2 reviews the multivalent interaction between sugar and proteins. Section 3 presents the phisycal chemical properties of glycopolymers. Section 4 presents the graft of glycopolymers and the biomaterial fabrication. Section 5 presents the glycopolymer in‐

The saccharide-protein interaction plays important roles in the living system, and the novel biomaterial fabriction is expected using the interaction. However, the saccharideprotein interaction is basically weak, and it is difficult to utilize and detect the interac‐ tions. It has been reported that the saccharide-protein interaction can be amplified by the multivalency [6, 7, 8]. Actually, saccharides on the cell-surfaces are displayed in a multi‐ valent manner. The glycolipids form densely saccharide structures of lipid-rafts [9], and glycoproteins usually have multivalent saccharide structures, which provies the multiva‐

The artificial multivalent saccharide displays also enables the multivalent interaction be‐ tween saccharide and protein. Various artificial compounds with multivalent saccharides have been reported (Figure 1). Proteis are commonly used as carries for the multivalent pre‐ sentation of antigens, and bovine serum albumin (BSA) is the representative [10]. Peptides are used as a scaffold of saccharide display [11]. Saccharide conjugates with DNA [12], cy‐ clodextrin [13] and polymers have been also reported to exhibit multivalent interactions.

the saccharide-protein interactions are also utilized for the biosensor of diseases.

terface with dendrimer.

Applications

456

**2. Multivalent interaction**

lent saccharide-protein interactions.

**Figure 1.** Schematic illustration of multivalent saccharide compounds.

There have been various multivalent saccharide derivatives as we described in the above section. Glycopolymers have been reported to exhibit larger multivalent effect comparing to other multivalent saccharides, because glycopolymers form large multivalent cluster [16]. The glycopolymers are the interesting compounds with large molecular weights and diverse structures. The glycopolymers are prepared by saccharide addition to polymer via polymer reaction, or by polymerization of saccharide monomers. The technique of synthetic polymer enables the preparation of versatile biomaterials. Especially, living radical polymerization is applicable to various saccharide monomers and provides the facile strategy for functional material preparation [17].

**Figure 2.** Chemical structure of monomers for glycopolymer preparation for (a) living radical polymerization, (b) ringopening metathesis polymerization and (c) polymerization with saccharide addition.

The various saccharide monomers have been reported, which were shown in Figure 2. There are various saccahride vinyl compounds. Styrene [18, 19], methacrylate [20], acrylate [21], acrylamide [22] and methacryl amide [23] with saccharide were reported. Living radical pol‐ ymerizations were reported with them. Norbornene saccharide derivatives were also report‐ ed, which provides the fine-tuned polymers via ring opening metastasis polymerization (ROMP) [24]. Reactive functional monomers were also utilized for glycopolymer synthesis. Monomers with acetylene [25] and active ester [26] were reported, where glycopolymers were obtained by polymerization and successive sugar addition.

pramolecular complex with various hydrophobic fluorophore and π-conjugate poymer of

Molecular Recognition of Glycopolymer Interface

http://dx.doi.org/10.5772/54156

459

Self-assembling properties of amphiphilic polymers were used in order to organize the gly‐ copolymer interface. The glycopolymer, PVLA, had amphiphilic structure and adsorbed to the hydrophobic interface [33]. PVLA adsorbed the hydrophobic polystyrene culture dish, and the culture dish was used as hepatocyte culture [34]. The adsorption of PVLA was in‐ vestigated with hydrophobic self-assembled monolayer (SAM) of octadecyltrimethoxysilane [35]. PVLA selectively adsorbed onto the hydrophobic substrate, exhibiting the lectin and hepatocyte affinity. We utilized the adsorption process to fabricate the micropatterned cell

On the other hand, the self-assembling properties were expanded to the complex and mi‐ cropatterned cell cultivation systems. We fabricated the micropatterned substrate with hydrophobic and cationic SAM The micropatterned substrates were fabricated by the for‐ mation of SAM and micropatterning with photolithography. The orthogonal self-assem‐ bly was performed with PVLA and anionic polysaccharide of heparin. PVLA and heparin bound to hydrophobic and cationic part, respectively. PVLA showed affinity to hepatocyte, and heparin binds to bFGF that has affinity to fibroblast cell. The multiple cell cultivation was accomplished with PVLA/hepatocyte and heparin/bFGF/fibroblast in

Glycopolymer-coated substrates were facilely prepared by self-assembly of hydrophobic in‐ teraction. However, it is difficult to control the density of glycopolymer by self-assembling

**Figure 3.** Chemical structure and properties of lactose-carrying polystyrene.

cultivation system and protein display.

a self-assembling manner [36].

**3. Grafted glycopolymers**

polythiophen [32].

Saccharide recognition proteins are called lectin, which basically have multiple domain structures [27]. The multivalent saccharides gain in enthalpy due to multiple binding to sug‐ ar recognition sites, and gain in entropy due to the various binding modes. The glycopoly‐ mers are large sugar cluster to gain the Gibbs free energy in both enthalpy and entropy, and lectins have multiple and valuable structure, which is advantage for binding. The distance of sugar binding sites is different with each lectin, which is easily tuned by copolymeriza‐ tion. The density, distance, and the size of multivalent compounds can be easily adjusted by copolymerization, which can be applied to variable lectins.

The glycopolymers are water soluble polymers, which can be utilized as artificial polymeric ligands or polymer drugs. Choi et al reported polyacrylic acid with sialic acid, and the poly‐ mer efficiently inhibited the sialidase of Influenza virus [6] Kobayashi et al reported the var‐ ious glycopolymers. The lactose substituted polystyrene (poly(*N*-vinylbenzyl-O-β-Dgalactopyranosyl-(1-4)-D-gluconamide (PVLA)) strongly interacted with lectin, and it was applied for hepatocyte culture [18]. Polystyrene with sialyl lactose showed the strong bind‐ ing to influenza virus A [28]. Gestwicki et al prepared various glycopolymers via metastasis reaction, and reported the glycopolymers to bind lectin and *E. coli* [29]. Nishimura reported the glycopolymers interacting with glycosyl transferases to synthesize oligosaccharides, and they developed the oligosaccharide synthesizer with glycopolymers [30].

### **2.2. Amphiphilic property of glycopolymer**

We defined glycopolymers as polymers with pendant saccharides. As we described above, the glycopolymers showed the strong multivalent effect based on the multivalency, with lec‐ tins, cells, viruses and bacteria. Another interesting property of glycopolymer is amphiphi‐ licity. Glycopolymers via addition polymerization have hydrophobic backbones with C-C bond, and are amphiphilic due to the hydrophilic side chain. The side chain of glycopoly‐ mer is bulky structure, and so the glycopolymer easily form the self-assembling structure. The structure of PVLA in aqueous solution was analyzed by small angle X-ray scattering [31], and it was found that PVLA formed rod-like structure, where the rod had the structure with long axis 10 nm and shot axis 5 nm.The rod-structure was similar to some polysacchar‐ ides of amylose and sizofiran.

It has been known that amylose is a host of hydrophobic compound as it is known starch-amylose complex. PVLA also became a host compound of hydrophobic substan‐ ces. We investigated the supramolecular polymer complex of PVLA. PVLA formed su‐ pramolecular complex with various hydrophobic fluorophore and π-conjugate poymer of polythiophen [32].

**Figure 3.** Chemical structure and properties of lactose-carrying polystyrene.

Self-assembling properties of amphiphilic polymers were used in order to organize the gly‐ copolymer interface. The glycopolymer, PVLA, had amphiphilic structure and adsorbed to the hydrophobic interface [33]. PVLA adsorbed the hydrophobic polystyrene culture dish, and the culture dish was used as hepatocyte culture [34]. The adsorption of PVLA was in‐ vestigated with hydrophobic self-assembled monolayer (SAM) of octadecyltrimethoxysilane [35]. PVLA selectively adsorbed onto the hydrophobic substrate, exhibiting the lectin and hepatocyte affinity. We utilized the adsorption process to fabricate the micropatterned cell cultivation system and protein display.

On the other hand, the self-assembling properties were expanded to the complex and mi‐ cropatterned cell cultivation systems. We fabricated the micropatterned substrate with hydrophobic and cationic SAM The micropatterned substrates were fabricated by the for‐ mation of SAM and micropatterning with photolithography. The orthogonal self-assem‐ bly was performed with PVLA and anionic polysaccharide of heparin. PVLA and heparin bound to hydrophobic and cationic part, respectively. PVLA showed affinity to hepatocyte, and heparin binds to bFGF that has affinity to fibroblast cell. The multiple cell cultivation was accomplished with PVLA/hepatocyte and heparin/bFGF/fibroblast in a self-assembling manner [36].
