Periplasmic-binding protein-based biosensors and bioanalytical assay platforms: Advances, considerations, and strategies for optimal utility

•Periplasmic binding proteins fulfill a biosensing niche for non-immunogenic targets.•Both homogeneous and heterogeneous assay platforms have been developed.•Conformational shifts upon binding require immobilization and labeling considerations.•Largely limited to natural targets, challenges remain in engineering for non-natural ligands.•PBP-based assays allow probing of microbiological nutrient response and communication. Periplasmic binding proteins provide gram-negative bacteria with mechanisms for nutrient uptake and sensing their environments through recognition and transport of small molecules and ions. In the assay development realm, the biosensing niche that these proteins fulfill is the recognition and detection of non-immunogenic targets, including inorganic ions, vitamins, amino acids, and sugars with affinities in the low nM to low μM range. These proteins have been applied in a variety of homogeneous and heterogeneous platforms including those based on enzyme-linked immunosorbent assays, surface plasmon resonance, immunomagnetic separation, and fluorescence resonance energy transfer. Their binding repertoire is largely limited to their natural targets, although progress has been made in engineering the target specificity by grafting alternate binding pockets and understanding the stability limitations inherent in engineering proteins for non-native targets. There has been significant interest in the glucose-galactose binding protein as an alternative to enzymes for diabetic glucose monitoring, but emerging applications for these proteins include environmental and microbiological sensing. Within, a comprehensive review of sensor advances is presented from a perspective of assay design, engineering challenges, and practical application. Immobilization strategies, labeling considerations, and overcoming stoichiometric signal limitations to yield optimal performance are discussed. [Display omitted]