Functional analysis of small-molecule and protein binding domains using monomeric streptavidin-based two-component biosensors
Protein biosensors are important analytical and diagnostic tools that convert molecular recognition into measurable activity. Most common two-component biosensors require two non-competing recognition domains, yet binding affinity and specificity alone do not reliably predict their compatibility with the rest of the biosensor. We report a modular platform that exploits monomeric streptavidin…
Protein biosensors are crucial tools for analyzing and diagnosing molecular interactions. Typically, these biosensors employ two non-competing recognition domains, although binding affinity and specificity do not always guarantee compatibility with the overall biosensor structure. Researchers have now developed a modular platform that utilizes monomeric streptavidin (mSA2) as a substitute recognition domain.
This innovation allows for quick functional assessment of potential binders within the biosensor framework, eliminating the need for an additional ligand-specific binding protein. The team established a chromatography-free method for extracting functional mSA2-based biosensor components from Escherichia coli bacteria. Through this approach, they successfully tested biotinylated small molecules, such as methotrexate and vitamin D3, as well as heart failure protein biomarkers galectin-3 and S100A7.
The results showed that the ligand-dependent reporter biosensor activation was successful. This newly developed platform offers a scalable and efficient solution for rapid binder evaluation and biosensor prototyping, significantly simplifying the process of biosensor development using binding domains generated through directed evolution or computational protein design.
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