Aptamers families

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. 

The data for specificity are equally surprising. As shown in Table 20.1, an aptamer aimed at basic fibroblast growth factor (bFGF) does not bind tightly to other members of the FGF family or to other proteins known to interact with an acidic substance, heparin (these data were reported in [5]). Similar data exist for aptamers aimed at several reverse transcriptases, serine proteases, P- and L-selectin, and cytokines such as VEGF and PDGF. Aptamers in vitro show extreme specificity for their intended targets. 

Where the identification of either DNA or RNA aptamers is required, alignment of either set of sequences post several rounds of screening should lead to the identification of critical consensus sequences, required for the desired aptameric activity. Often this results in families of aptamers - competition experiments may then be performed to see whether these families... 

Scheme 5. Secondary structure on Escherichia coli 16S RNA in the region of decoding A-site 20 and its shortened hairpin version (27-mer model oligoribonucleotide 21) used for the structure determination with bound paromomycin (16) [125]. Nucleotides which are protected from chemical modification by bound aminoglycosides, methylated in resistant strains or essential for aminoglycoside binding are shown in bold. For more detailed discussion refer to text. The sequence of the 27-mer RNA aptamer 22 with its secondary structure is shown. The consensus sequence identified for this family of aptamers is highlighted in bold [141].

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