Molecular dynamic simulation of FimD ssDNA aptamers and development of SPR aptasensor for Salmonella

For more than a century, non-typhoidal Salmonella has been recognised as a major foodborne pathogen, responsible for human illness underscoring its continued importance to public health. In the current study, we employed molecular dynamic simulation and Surface Plasmon Resonance (SPR) biosensing based experimental approaches to assess the interaction between the ssDNA aptamers (DFRM13, DFRM110, DFRM29, DFRM33, and DFRM39) and Salmonella FimD usher protein. The docking studies revealed superior binding performance for DFRM13-FimD complex based on its low RMSD (0.7 ± 0.4 nm) and z-score (-1.9) values followed by DFRM39-FimD complex (RMSD: 1.1 ± 0.6 nm, z-score: -1.6). The RMSD, RMSF, Rg, SASA analysis and high number of H-bonds with the ssDNA aptamer (∼18 bonds) observed during 50 ns molecular dynamics simulations revealed that DFRM29-FimD complex also exhibited stable binding interactions. SPR assay optimized with biotinylated aptamers on streptavidin sensors exhibited relatively lower dissociation constant (K_D) or higher affinity for native FimD protein from Salmonella lysates (K_D10^-7 to 10^-8 M) than rFimD (K_D10^-6 to 10^-7 M) due to the cross-over SELEX approach adopted for enriching the aptamers. The DFRM13 demonstrated lowest K_D toward FimD in Salmonella thermal lysates (K_D 1.24 × 10^-8 M), followed by DFRM39 (K_D 4.17 × 10^-8 M) indicating highest binding affinity among other aptamers as determined by SPR interaction kinetics. The sensorgrams with curvatures characteristic of FimD detection was not observed when thermal lysates of non-Salmonella bacteria were examined in the SPR assay, thus confirming the specificity of aptamers for Salmonella detection. Overall, the computational and experimental findings from our study demonstrated potential for possible utilization of the aptamers DFRM13 and DFRM39 in Salmonella diagnostic platforms.