Specific glycosidase activity isolated from a random phage display antibody library

Carbohydrates serve as key receptor sites in various cellular events such a s viral attachment, tumor formation, and tissue inflammation. A potential r oute to control these events is to manipulate targeted carbohydrate structu res in vivo using specifically designed glycohydrolases. Here we show that a stereospecific catalytic activity designed toward a particular sugar and linkage can be readily isolated from a phage display antibody library deriv ed from a nonimmunized host. The activity was isolated using a transition-s tate analogue mimicking an alpha -glucosidasic linkage as antigen and showe d a 20-fold specificity for that sugar and linkage. The DNA sequence, howev er, contains a large deletion in the antibody gene, which also changes the downstream reading frame, resulting in a translated sequence containing onl y 57 amino acids that has a predominantly hydrophobic amino terminal and a strongly hydrophilic carboxy terminal. The isolated catalytic activity has a strong pH dependence, attributable to one or more of the numerous potenti ally charged groups in the carboxyl terminal. While the protein readily for ms more stable multimers, the 7.3-kD monomer represents by far the smallest glycosidase enzyme reported to date and can provide substantial new inform ation toward understanding and modifying glycosidase activity.