Influence of Precise Polymer Conjugation on Aptamer-Target Binding

The effect of polymer bioconjugation on the binding properties of the ATP aptamer was studied using bio-layer interferometry (BLI) and surface plasmon resonance (SPR). For this study, ten different polymer-aptamer bio-hybrids were synthesized by automated solid-phase phosphoramidite chemistry. These bio-hybrid macromolecules contain <i>(i)</i> a DNA segment (<i>i.e.</i>, either the ATP aptamer or a control sequence with no ATP affinity), <i>(ii)</i> one or two synthetic poly(phosphodiester) segments containing either propyl, triethylene glycol, or pentaethylene glycol spacers, and <i>(iii)</i> a biotin end-group allowing immobilization on streptavidin sensors for BLI and SPR experiments. Diblock and triblock architectures were prepared in order to assess the influence of the number of conjugated polymer chains on aptamer-target binding. All bio-hybrid polymers were characterized by high resolution electrospray mass spectrometry, ion-exchange HPLC, and polyacrylamide gel electrophoresis. All these methods confirmed the formation of the targeted bio-hybrids. Furthermore, BLI and SPR experiments demonstrated that all bio-hybrid macromolecules containing the ATP aptamer sequence could bind ATP, indicating that polymer conjugation did not compromise the aptamer's functionality, even when positioned between two synthetic chains.