Screening for Activity

On the assumption that the library will hold some sequences that will be tailored for activity against particular proteases of interest, the task is to find which library component has activity. Activity is characterised by tight non-covalent binding, and so the enzyme itself can be used as a probe to discover which members of the library show activity. It is possible to tag the protease enzyme in such a way that the presence of this protein is characterised by a colour reaction. Addition of the protein to a suspension of the library will then cause any resin beads that bind the protein to be coloured dark blue. These beads can be easily visualised under a low power microscope, and separated from the non-binding sequences.

Under the microscope, beads holding sequences that bind tightly to the protease enzyme become highly stained. In the actual micrograph shown above there is a single active bead (center) amongst a large number of inactive beads. The scale bar is 0.2 mm. Approximately 1 in 1000 of the library members were found to show activity, illustrating the selectivity of the interaction and the power of library techniques to identify active components.