ABSTRACT
Staphylococcus aureus (S. aureus), a common foodborne pathogen, poses significant public health challenges due to its association with various infectious diseases. A key player in its pathogenicity, which is the IsdA protein, is an essential virulence factor in S. aureus infections. In this work, we present an integrated in-silico and experimental approach using MD simulations and surface plasmon resonance (SPR)-based aptasensing measurements to investigate S. aureus biorecognition via IsdA surface protein binding. SPR, a powerful real-time and label-free technique, was utilized to characterize interaction dynamics between the aptamer and IsdA protein, and MD simulations was used to characterize the stable and dynamic binding regions. By characterizing and optimizing pivotal parameters such as aptamer concentration and buffer conditions, we determined the aptamer's binding performance. Under optimal conditions of pH 7.4 and 150 mM NaCl concentration, the kinetic parameters were determined; ka = 3.789 × 104/Ms, kd = 1.798 × 103/s, and KD = 4.745 × 10-8 M. The simulations revealed regions of interest in the IsdA-aptamer complex. Region I, which includes interactions between amino acid residues H106 and R107 and nucleotide residues 9G, 10U, 11G and 12U of the aptamer, had the strongest interaction, based on ΔG and B-factor values, and hence contributed the most to the stability of the interaction. Region II, which covers residue 37A reflects the dynamic nature of the interaction due to frequent contacts. The approach presents a rigorous characterization of aptamer-IsdA binding behavior, supporting the potential application of the IsdA-binding aptamer system for S. aureus biosensing.
ABSTRACT
Bioaffinity nanoprobes are a type of biosensor that utilize the specific binding properties of biological molecules, such as antibodies, enzymes, and nucleic acids, for the detection of foodborne pathogens. These probes serve as nanosensors and can provide highly specific and sensitive detection of pathogens in food samples, making them an attractive option for food safety testing. The advantages of bioaffinity nanoprobes include their ability to detect low levels of pathogens, rapid analysis time, and cost-effectiveness. However, limitations include the need for specialized equipment and the potential for cross-reactivity with other biological molecules. Current research efforts focus on optimizing the performance of bioaffinity probes and expanding their application in the food industry. This article discusses relevant analytical methods, such as surface plasmon resonance (SPR) analysis, Fluorescence Resonance Energy Transfer (FRET) measurements, circular dichroism, and flow cytometry, that are used to evaluate the efficacy of bioaffinity nanoprobes. Additionally, it discusses advances in the development and application of biosensors in monitoring foodborne pathogens.
