Supramolecular tandem enzyme assays for multiparameter sensor arrays and enantiomeric excess determination of amino acids.
The coupling of an enzymatic transformation with dynamic host-guest exchange allows the unselective binding of macrocycles to be used for highly selective analyte sensing. The resulting supramolecular tandem enzyme assays require the enzymatic substrate and its corresponding product to differ significantly in their affinity for macrocycles, for example, cation receptors, and to show a differential propensity to displace a fluorescent dye from its host-guest complex. The enzymatic transformation results in a concomitant dye displacement that can be accurately followed by optical spectroscopy, specifically fluorescence. By exploiting this label-free continuous enzyme assay principle with the fluorescent dye Dapoxyl and the macrocyclic host cucurbit[7]uril, a multiparameter sensor array has been designed, which is capable of detecting the presence of amino acids (e.g. histidine, arginine, lysine, and tyrosine) and their decarboxylases. Only in the presence of both, the particular amino acid and the corresponding decarboxylase, is the amine or diamine product formed. These products are more highly positively charged than the substrate, have a higher affinity for the macrocycle and, therefore, displace the dye from the complex. The extension of the high selectivity and muM sensitivity of the tandem assay principle has also allowed for the accurate measurement of D-lysine enantiomeric excesses of up to 99.98 %, as only the L-enantiomer is accepted by the enzyme as a substrate and is converted to the product that is responsible for the observed fluorescence signal.
Bailey
David M
DM
School of Engineering and Science, Jacobs University Bremen, Campus Ring 1, 28759 Bremen, Germany.
Hennig
Andreas
A
Uzunova
Vanya D
VD
Nau
Werner M
WM
D000596:Amino Acids / Q000737:chemistry* / Q000378:metabolism*; D002151:Calorimetry; D004798:Enzymes / Q000032:analysis* / Q000378:metabolism*; D015394:Molecular Structure; D013237:Stereoisomerism; D013816:Thermodynamics; D017186:Titrimetry
D000596:Amino Acids; D004798:Enzymes