Spacer modifications

Modes of attachment of functional groups to crosslinked polystyrene are discussed ( 1). Attention is drawn to improved stability and activity of polymer-bound reagents and catalysts incorporating dimethylene spacer between polystyrene aryl and functional group heteroatom, and the simplicity and versatility of their synthesis through high-conversion functional group modifications. 

H. Sashiwa, Y. Shigemasa, and R. Roy, Chemical modification of chitosan. 3. Hyperbranched chitosan-sialic acid dendrimer hybrid with tetraethylene glycol spacer, Macromolecules, 33 (2000) 6913-6915. 

Since sialic acid is a frequent terminal sugar constituent of the polysaccharide trees on glycoproteins, this method selectively forms reactive aldehydes on the most accessible parts for subsequent modifications. The carbohydrate polymer of a protein provides a long spacer arm that can be used to conjugate another large macromolecule, such as a second protein, with little steric problems. 

A common choice of crosslinker for this type of reaction is sulfo-SMCC, which has been used extensively for antibody conjugation (Chapter 20, Section 1.1). A better option for dendrimer conjugation is to use a similar crosslinker design, but one that contains a hydrophilic PEG spacer arm to promote dendrimer hydrophilicity after modification. Derivatization of an amine-dendrimer with a NHS-PEG-maleimide can create an intermediate that is coated with water-soluble PEG spacers. This modification helps to mask any potential for nonspecific interactions that the PAMAM surface may have, while providing terminal thiol-reactive maleimides for coupling ligands (Figure 7.10). 

Hong et al. (2004) also found that modification of PAMAM dendrimers with a short PEG linker arm could act to reduce nonspecificity caused by the amines on the dendrimer-modified surface. An azido-PEGj-aininc spacer was activated with nitrophenyl carbamate to yield an activated intermediate that could be used to modify the amines on the dendrimer (Figure 7.24). Reaction at high molar ratio resulted in about 61 PEG-azido spacers on the dendrimer. Reduction of the azido group to an amine using triphenylphosphine in THF provided the dendrimer-PEG-amine derivative for surface modification. The added presence of the PEG spacer arm reduced... 

Although NHS-LC-biotin and sulfo-NHS-LC-biotin are very popular reagents for biotinylation, they both result in hydrophobic aliphatic biotin modifications on proteins and antibodies. Unfortunately, these groups have a tendency to aggregate in aqueous solution and may cause protein precipitation or loss of activity over time. For this reason, the use of more hydrophilic PEG-based biotin compounds of approximately the same spacer length may be a better alternative for maintaining water solubility of modified proteins (Chapter 18). 

Figure 13.4 APTS-modified surfaces may be further derivatized with amine-reactive crosslinkers to create additional surface characteristics and reactivity. Modification with NHS-PEG4-azide forms a hydrophilic PEG spacer terminating in an azido group that can be used in a click chemistry or Staudinger ligation reaction to couple other molecules.

Figure 14.11 The modification of amine-containing particles with NHS-PEG4-maleimide produces hydrophilic PEG spacers containing terminal thiol-reactive groups. Coupling of thiol-containing proteins then results in the formation of thioether linkages.

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