Biotin receptors

Upon occupation of biotin receptors, the cell membrane becomes hyperpolarized. This hyperpolarization causes an increase in the posteriorly directed beating of the cell s propulsive cilia, and the cells move smoothly up the concentration gradient. Importantly, the hyperpolarization also decreases the likelihood of membrane depolarization, and if mild, slows the ciliary beat frequency and slows the cells, while if large, brings about a calcium action potential that reverses the ciliary... 

Fig. 10.9 pH-dependent biotin availability for binding, (a) Above pH 7.0, biotin anchored on the micelle core via pH-sensitive PHis is shielded by PEG shell of the micelle, (b) Biotin is exposed on the micelle surface at acidic conditions (6.5 < pH < 7.0) and can interact with cells, which facilitates biotin-receptor-mediated endocytosis. When the pH is further lowered (pH < 6.5), the micelle destabilizes, which enhances drug release, (modified and reproduced from [154], with permission from American Chemical Society)... 

To bring the nanocontainer to a specific place where it should release its pay-load, targeting is a required approach. Hence, much work has been carried out to attach ligands or antibodies to the hydroxyl end-group of PEG-based assemblies [150,181,243], Biotinylated nondegradable block copolymer assemblies have been shown to attach to surfaces coated with the biotin receptor avidin [146,147, 150,244], Coupling chemistry has been applied to conjugate either an antihuman IgG, or antihuman serum to PEG-carbonate- or PEG-polyester-assembled polymer vesicles [149,245], HIV-derived Tat peptide attached to PEG-PBD polymersomes enhanced the cellular delivery of nanoparticles [246] and increased dendritic cell uptake in vitro [181]. 

In an extensive SFA study of protein receptor-ligand interactions, Leckband and co-workers [114] showed the importance of electrostatic, dispersion, steric, and hydrophobic forces in mediating the strong streptavidin-biotin interaction. Israelachvili and co-workers [66, 115] have measured the Hamaker constant for the dispersion interaction between phospholipid bilayers and find A = 7.5 1.5 X 10 erg in water. 

A typical force curve showing the specific avidin-biotin interaction is depicted in figure Bl.20.10. The SFA revealed the strong influence of hydration forces and membrane undulation forces on the specific binding of proteins to membrane-bound receptors [81]. 

Direct measurement of the interaction potential between tethered ligand (biotin) and receptor (streptavidin) have been reported by Wong et al [16] and demonstrate the possibility of controlling range and dynamics of specific biologic interactions via a flexible PEG-tether. 

Patterning of enzyme monolayers on a solid surface was carried out by photoactivation of immobilized monolayer of caged -biotin derivatives in selected areas. Specific oriented binding of enzyme-avidin conjugates could be readily made to the photoactivated zones [42]. Oriented immobilization of G-protein-coupled receptors on a solid surface was also made possible on a biotinylated surface by first immobilizing streptavidin, followed by the immobilization of biotinylated G-protein-coupled receptor [43]. 

Fig. 10. N4 ligand systems for conjugation 99mTc to biomolecules R = in vivo targeting biomolecules (e.g., biotin [100, 101], somatostatin receptor-avid peptide [103,104])...

In another example, ligands can be biotinylated with a cleavable biotinylation reagent and then incubated with receptor molecules. The resulting complex can be isolated by affinity chromatography on immobilized (strept)avidin. Final purification of the ligand-receptor can be accomplished by cleaving the biotin modification sites while the complex is still bound to the support. The receptor complex thus can be eluted from the column without the usual harsh conditions required to break the avidin-biotin interaction. 

NHS esters of D-biotin have been used in many applications, including the biotinylation of rat IgE to study receptors on murine lymphocytes (Lee and Conrad, 1984), in the development of... 

Using a similar approach, Clq has been modified with biotin-HPDP and allowed to interact with its specific receptor. Subsequent purification of the Clq receptor was accomplished through cleavage of the disulfide bridge of the biotinylation reagent (Ghebrehiwet et al., 1988). 

Biotin-hydrazide has been used to biotinylate antibodies at their oxidized carbohydrate residues (O Shanessy et al., 1984, 1987 O Shanessy and Quarles, 1985 Hoffman and O Shannessy, 1988), to modify the low-density lipoprotein (LDL) receptor (Wade et al., 1985), to biotinylate nerve growth factor (NGF) (Rosenberg et al., 1986), and to modify cytosine groups in oligonucleotides to produce probes suitable for hybridization assays (Reisfeld et al., 1987) (Chapter 27, Section 2.3). 

Wade, D.P., Knight, B.F., and Soutar, A.K. (1985) Detection of the low-density lipoprotein receptor with biotin-low-density lipoprotein. A rapid new method for ligand blotting. Biochem. J. 229, 785-790. 

Immunosensors have been developed commercially mostly for medical purposes but would appear to have considerable potential for food analysis. The Pharmacia company has developed an optical biosensor, which is a fully automated continuous-flow system which exploits the phenomenon of surface plasmon resonance (SPR) to detect and measure biomolecular interactions. The technique has been validated for determination of folic acid and biotin in fortified foods (Indyk, 2000 Bostrom and Lindeberg, 2000), and more recently for vitamin Bi2. This type of technique has great potential for application to a wide range of food additives but its advance will be linked to the availability of specific antibodies or other receptors for the various additives. It should be possible to analyse a whole range of additives by multi-channel continuous flow systems with further miniaturisation. 

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