High-affinity aptamers selectively inhibit human nonpancreatic secretory phospholipase A2 (hnps-PLA2)

A family of sequence-related 2'-aminopyrimidine, 2'-hydroxylpurine aptamers, developed by oligonucleotide-based combinatorial chemistry, SELEX (systematic evolution of ligand by exponential enrichment) technology, binds human nonpancreatic secretory phospholipase A(2) (hnps-PLA(2)) with nanomolar affinities and inhibits enzymatic activity. Aptamer 15, derived from the family, binds hnps-PLA(2) with a K-d equal to 1.7 +/- 0.2 nM and, in a standard chromogenic assay of enzymatic activity, inhibits hnps-PLA(2) with an IC50 of 4 nM, at a mole fraction of substrate concentration of 4 x 10(-6) and a calculated K-i of 0.14 nM. Aptamer 15 is selective for hnps-PLA(2), having a 25- and 2500-fold lower affinity, respectively, for the unrelated proteins human neutrophil elastase and human IgG. Contractions of guinea pig lung pleural strips induced by hnps-PLA(2) are abolished by 0.3 mu M aptamer 15, whereas contractions induced by arachidonic acid are not altered. The structure that is essential for binding and inhibition appears to be a 40-base hairpin/loop motif with an asymmetrical internal loop. The affinity and activity of the aptamers demonstrate the ability of the SELEX process to isolate antagonists of nonnucleic-acid-binding proteins from vast oligonucleotide combinatorial libraries.