Biotin ligand

Avidin is a glycoprotein that is isolated from egg white. In its active form, it exists as a tetramer which is composed of four identical subunits that are held together by noncovalent interactions. This tetramer, which has a molecular weight of 64 kDa, binds four biotin ligands (each with a molecular weight of 244 Da). Four biotin molecules bind the tetrameric avidin at sites that are partially created by the quaternary structure of avidin. The avidin-biotin complex is one of the strongest known non-covalent com-... 

Fig. 2 Biotin-avidin technology Artificial metalloenzymes [M(L )(biotin-ligand)]c(strept)avidin for enantioselective catalysis are based on the anchoring of a catalyticaUy active metal fragment within a host protein via a hgand, a spacer, and biotin. Chemical optimization can be achieved either by varying the spacer or the metal chelate moiety ML ). Saturation mutagenesis at a position close to the metal moiety ( ) can be used for genetic optimization...

A water soluble biotin ligand is now available from several vendors that may alleviate the solubility and handling problems of the currently described biotin ligand. However, its efficacy in this protocol is yet to be determined. 

Figure 15 Schematic representation of the procedure used for patterning biotin ligands onto SAMs consisting of activated carboxylic esters. (From J. Lahiri, E. Ostuni, and G.M. Whitesides. Langmuir 15 2055-2060, 1999. With permission.)...

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. 

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. 

The avidin-biotin complex, known for its extremely high affinity (Green, 1975), has been studied experimentally more extensively than most other protein-ligand systems. The adhesion forces between avidin and biotin have been measured directly by AFM experiments (Florin et al., 1994 Moy et al., 1994b Moy et al., 1994a). SMD simulations were performed on the entire tetramer of avidin with four biotins bound to investigate the microscopic detail of nnbinding of biotin from avidin (Izrailev et al., 1997). 

Grubmiiller et al., 1996] Grubmiiller, H., Heymann, B., and Tavan, P. Ligand binding and molecular mechanics calculation of the streptavidin-biotin rupture force. Science. 271 (1996) 997-999... 

Both the AFM rupture experiments as well as our simulation studies focussed on the streptavidin-biotin complex as a model system for specific ligand binding. Streptavidin is a particularly well-studied protein and binds its ligand biotin with high affinity and specificity [51]. Whereas previous experiments (see references in Ref. [49]) and simulation studies [52] referred only to bound/unbound states and the associated kinetics, the recent AFM... 

Thus, we have found unexpected complexities and even in this simple system have not yet been unable to accurately extrapolate the results of simulations done over periods varying from 1 to several hundred ps, to the low-friction conditions of extraction experiments performed in times on the oi dc r of ms. The present results indicate that one should not expect agreement between extraction experiments and simulations in more complex situations typically found in experiments, involving also a reverse flow of water molecules to fill the site being evacuated by the ligand, unless the simulation times are prolonged well beyond the scope of current computational resources, and thereby strengthen the conclusion reached in the second theoretical study of extraction of biotin from it.s complex with avidin [19]. 

Using a combination of techniques such as EPR, resonance Raman, and MCD spectroscopy, the conversion of [2Fe-2S] into [4Fe—4S] centers has been found to take place under reducing conditions in E. coli biotin synthase 281). The as-prepared form of this enzyme has been thought to contain one [2Fe-2S] center per monomer, coordinated by the three cysteine residues of the motif Cys-X3-Cys-X2-Cys and by a fourth, noncysteinyl ligand. Upon reduction, a [4Fe-4S] cluster bridging two monomers may be formed in the active enzyme. In the reduced state, the [4Fe-4S] center is characterized by a mixture of S = I and S = k spin states giving EPR features at g 5.6 and... 

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])...

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