A homogeneous fluorometric assay platform based on novel synthetic proteins.
Novel synthetic recombinant sensor proteins have been created to detect analytes in solution, in a rapid single-step "mix and read" noncompetitive homogeneous assay process, based on modulating the Förster resonance energy transfer (FRET) property of the sensor proteins upon binding to their targets. The sensor proteins comprise a protein scaffold that incorporates a specific target-capturing element, sandwiched by genetic fusion between two molecules that form a FRET pair. The utility of the sensor proteins was demonstrated via three examples, for detecting an anti-biotin Fab antibody, a His-tagged recombinant protein, and an anti-FLAG peptide antibody, respectively, all done directly in solution. The diversity of sensor-target interactions that we have demonstrated in this study points to a potentially universal applicability of the biosensing concept. The possibilities for integrating a variety of target-capturing elements with a common sensor scaffold predict a broad range of practical applications.
Vardar-Schara
Gönül
G
Department of Molecular Biosciences and Bioengineering, University of Hawaii, Honolulu, HI 96822, USA.
Krab
Ivo M
IM
Yi
Guohua
G
Su
Wei Wen
WW
D001426:Bacterial Proteins / Q000235:genetics; D015374:Biosensing Techniques; D002352:Carrier Proteins / Q000235:genetics; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D005456:Fluorescent Dyes / Q000737:chemistry*; D049452:Green Fluorescent Proteins / Q000235:genetics; D008164:Luminescent Proteins / Q000737:chemistry* / Q000235:genetics; D058619:Maltose-Binding Proteins; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000737:chemistry
D001426:Bacterial Proteins; D002352:Carrier Proteins; C485182:Cyan Fluorescent Protein; D005456:Fluorescent Dyes; D008164:Luminescent Proteins; D058619:Maltose-Binding Proteins; D011993:Recombinant Fusion Proteins; C054562:yellow fluorescent protein, Bacteria; D049452:Green Fluorescent Proteins