Protein kinase family binding assays

Third, we have selected aptamers that can bind to the 3II isozyme of protein kinase C from an RNA pool that spanned 120 random positions [5], The aptamers fell into several families, and individuals from two of the most prominent families were assayed for their ability to inhibit the enzymatic activity ofPKC. While these aptamers efficiently inhibited the enzymatic activity of the 3II isozyme, they had a 10-fold lower K, for the 3I isozyme (96% similar) and showed no activity against the a isozyme (80% similar). These specificities rival those seen with monoclonal antibodies. We have now selected aptamers that can bind to the a isozyme ofPKC from the same RNA pool. Sequence comparisons of the anti-pil and anti-a aptamers (Fig. 13) suggest that the map that relates target space and sequence space is convoluted. For example, while one family of aptamers was returned from both selections, other families were unique for one or the other isozyme. 

By combining assays from the nine providers in Appendix 1.1, inhibitor binding for up to 407 wild type protein or lipid kinases can be assessed. The data in Appendix 1.1 shows that the kinase families can be well covered by a combination of the commercial assay panels. Where one panel may be deficient, other panels provide coverage. 

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