Boronic acid-salicylhydroxamate reagent pairs

The interaction of PBA derivatives with molecular species having the above functional groups occurs optimally in the pH range of 8-9, but it is typically reversible at acid pH or in the presence of a high concentration of competing ligand. However, the heterocyclic boronic acid complex is relatively stable under optimal conditions of formation. 

Complex formation between PBA or PDBA group and the SHA group occurs in a wide range of buffer types from pH 5 to 9, and it can tolerate high salt conditions (to 1.5 M) or the 

Bergseid et al. (2000) also reported on the use of SHA-activated chromatography supports for the coupling of boronate-containing affinity ligands. In this case, immobilized RNase A was used to purify anti-RNase antibodies from antiserum samples. RNase A was modified with an NHS-PDBA crosslinker at a molar ratio of 100 1 (crosslinkenprotein), purified to remove excess crosslinker, and then coupled to an SHA-agarose support in 0.1 M sodium bicarbonate, pH 8.0. 

The following protocol describes the immobilization of a protein on an amine-surface using the P(D)BA-SHA chemistry. 

Dissolve a first protein to be modified at a concentration of l-10mg/ml in 0.1 M sodium bicarbonate buffer, pH 8.5. PBS buffer at physiological pH also may be used as the reaction medium. 

---