Mannose-decorated cyclodextrin vesicles: The interplay of multivalency and surface density in lectin–carbohydrate recognition

Mannose-decorated cyclodextrin vesicles: The interplay of multivalency and surface density in lectin–carbohydrate recognition Ulrike Kauscher and Bart Jan Ravoo* Full Research Paper Address: Organic Chemistry Institute, Westfälische Wilhelms-Universität Münster, Correnstraße 40, 48149 Münster, Germany Email: Bart Jan Ravoo* - * Corresponding author Keywords: carbohydrates; cyclodextrins; lectins; molecular recognition; multivalency; vesicles Open Access Beilstein J. Org. Chem. 2012, 8, 1543–1551. doi:10.3762/bjoc.8.175 Received: 25 May 2012 Accepted: 17 August 2012 Published: 17 September 2012 This article is part of the Thematic Series "Superstructures with cyclodextrins: Chemistry and applications". Guest Editor: H. Ritter © 2012 Kauscher and Ravoo; licensee Beilstein-Institut. License and terms: see end of document. Abstract Cyclodextrin vesicles are versatile models for biological cell membranes since they provide a bilayer membrane that can easily be modified by host–guest interactions with functional guest molecules. In this article, we investigate the multivalent interaction of the lectin concanavalin A (ConA) with cyclodextrin vesicles decorated with mannose–adamantane conjugates with one, two or three adamantane units as well as one or two mannose units. The carbohydrate–lectin interaction in this artificial, self-assembled glycocalyx was monitored in an agglutination assay by the increase of optical density at 400 nm. It was found that there is a close relation between the carbohydrate density at the cyclodextrin vesicle surface and the multivalent interaction with ConA, and the most efficient interaction (i.e., fastest agglutination at lowest concentration) was observed for mannose–adamantane conjugates, in which both the cyclodextrin–adamantane and the lectin–mannose interaction is inherently multivalent. Introduction The surface modification of materials with carbohydrates has attracted much attention due to the fact that such materials can be compared to and compatible with the cell surface [1]. The “glycocalyx” is a dense layer on the surface of the cell, which serves as a responsive interface with its environment and also serves as a natural protective shield. The glycocalyx consists of various numbers and arrangements of polysaccharides and is found in eukaryotic as well as in prokaryotic cells. A wellknown example of the pivotal role of oligosaccharides on cell surfaces is the fact that human blood types (A, B, AB and 0) are solely determined by minor changes in the composition of the erythrocyte glycocalyx. Additionally, many biological mecha- 1543 Beilstein J. Org. Chem. 2012, 8, 1543–1551. nisms are mediated by multivalent recognition of carbohydrates. For example, lectins are proteins that bind to specific carbohydrates on the cell surface and activate biochemical responses [2]. In this way, protein–carbohydrate interactions regulate cell division, protein synthesis, the immune system, and the adhesion of cells. A well-known lectin is concanavalin A (ConA), which can be readily obtained from jack-beans. It has four identical binding sides and binds α-mannose, α-glucose and their derivatives. Because of the importance of carbohydrates and their multivalent recognition by lectins in physiological processes, they are also considered a promising tool for the development of drug-delivery systems [3]. Synthetic bilayer vesicles are a versatile model for biological cell membranes, and there are a substantial number of reports on synthetic glycolipids that mimic the glycocalyx [4-23]. Multivalent guest interaction with the surface of the vesicles has become a useful system to investigate recognition, adhesion and fusion of biological cell membranes [24-26]. In this context, amphiphilic cyclodextrins are a promising platform due to their ability to form stable bilayer vesicles that can be functionalized by self-assembly [27]. To this end, cyclodextrins are modified with long alkyl chains (“tails”) and short oligo(ethylene glycol) head groups. These macrocyclic amphiphiles form unilamellar bilayer vesicles in aqueous solution upon hydration of a thin film cast by evaporation from organic solution and extrusion through a 0.1 μm polycarbonate membrane [27]. The cavities of each cyclodextrin are available to form inclusion complexes with hydrophobic guest molecules. Adamantane is known to be an excellent guest for β-cyclodextrin cavities (Ka = (2–3) × 104 M−1). We were able to recently demonstrate the interaction of monovalent bifunctional guest molecules, containing a maltose or lactose unit and an adamantane unit, with cyclodextrin vesicles, and their ability to agglutinate with lectins [28]. We also showed that agglutination requires a critical density of carbohydrate ligand on the cyclodextrin vesicle surface [29]. In this work we investigate the influence of multivalent recognition by guest molecules with an increasing number of adamantane and mannose units. It is our hypothesis that more adamantane units in the guest molecule lead to higher affinity for the cyclodextrin vesicles due to multivalent interaction at the vesicle surface. In addition, we increased the number of mannose units in the guest molecule, assuming that a high density of carbohydrate is essential for multivalent lectin binding at the vesicle surface. Additionally, all guest molecules possess α-mannose units, which bind to lectins such as concanavalin A (ConA, Figure 1). Guest 1 contains a single mannose and a single adamantane unit. Guest 2 and guest 3 contain a single mannose and two or three adamantane units, respectively. Guest 4 contains two mannose as well as two adamantane units. The synthesis of 1–4 is described in Supporting Information File 1. The analytical data for 1–4 are fully consistent with their molecular structure. The synthesis of amphiphilic β-cyclodextrin 5 has been reported previously [30]. Unilamellar vesicles with a diameter of 100–150 nm are obtained by extrusion [27,30]. To investigate the ability of adamantane functions to bind into the cavity of cyclodextrins, the synthesized guest molecules were investigated regarding their 1:1 complexation behavior towards β-cyclodextrin. Isothermal titration calorimetry (ITC) was carried out with β-cyclodextrin and each of the synthesized guest molecules 1–4. The concentrations were chosen to provide one cyclodextrin cavity for each adamantane unit and are displayed in Table 1. The effective adamantane concentration describes the concentration of adamantane units. A guest with two adamantane units (2 or 4) results in an effective adamantane concentration that is twice the concentration of the divalent guest molecule. For guest 3, the effective adamantane concentration is three times the concentration of the trivalent guest molecule. The results of these titrations can be seen in Table 1 and Figure 2. Results and Discussion The thermodynamic parameters of guests 1–4 are characteristic of the formation of a 1:1 (...truncated)