In silico maturation of DNA aptamer against the prostate-specific antigen (PSA) and kinetic analysis

The detection of the prostate-specific antigen (PSA) serves as a critical marker for the diagnosis and follow-up of prostate cancer. DNA aptamers targeting PSA have been successfully screened using the systematic evolution of ligands by exponential enrichment (SELEX) technique, complemented by in silico maturation processes. In this study, we aim to optimize a truncated aptamer, denoted as TA87, through computational methods and to analyze potential aptamer candidates in the aptamer-PSA interactions. The PSA antibody, aptamer ΔPSap4#5, and an identified but unpublished aptamer, PSAG221, were evaluated in quartz crystal microbalance (QCM) experiments alongside aptamers derived from TA87. The Tanimoto similarity score and the ZDOCK program, coupled with the ZRANK scoring function, were adopted to assess the secondary structure of single-point mutants of TA87 and their binding interactions with PSA, respectively. Detailed analyses of the aptamer-protein complexes were conducted using molecular dynamics (MD) simulations. Mutations TA87M24 and TA87M49, along with PSAG221 and TA87, showed superior ZDOCK scores compared to ΔPSap4#5. MD simulations further suggested that PSAG221 aptamer might offer enhanced binding to PSA over ΔPSap4#5. The affinity constant (K_D) values for the antibody, ΔPSap4#5, PSAG221, TA87, TA87M24, and TA87M49 with PSA were determined through QCM measurements to be 0.35, 0.33, 0.35, 0.56, 0.45, and 0.51 μM^-1, respectively. The experimental results showed that the truncated aptamers, TA87, and the two mutations, TA87M24 and TA87M49, did not demonstrate superior PSA binding affinity. Aptamer PSAG221 demonstrated performance comparable to that of the antibody, although slightly inferior to ΔPSap4#5. The aptamer PSAG221 reported in this study could be an alternative probe for developing future PSA aptasensor platforms.