Structure-affinity balance of anti-cardiac troponin I aptamer: Effects of sequence truncation

The paper describes the rational design and truncation of high-affinity DNA aptamer ApTnI12 targeting cardiac troponin I (cTnI), a key biomarker for early diagnosis of cardiac injury. By combining molecular modeling, surface plasmon resonance, and bioluminescent assay, a series of truncated variants of ApTnI12 were obtained and studied to elucidate the role of its structural elements in mediating the aptamer's affinity and specificity. The G-quadruplex motif was shown to be essential for maintaining the aptamer stability and to play a key role in the cTnI epitope recognition, during which the flanking duplex regions and loops significantly contributed to binding strength and specificity through electrostatic and hydrogen bonding interactions. The specificity of the aptamer consisting of only G-quadruplex was significantly reduced, highlighting the need for careful monitoring of its non-specific binding. A rationally designed random duplex aptamer variant achieved comparable affinity to SELEX-derived aptamers, though with a compromise in specificity. These findings emphasize the importance of balancing structural elements in the aptamer design and underscore the need for integrated computational and empirical approaches for developing aptamers with optimal performance in biosensing and therapeutic applications.