Title of article :
Open tubular columns containing the immobilized ligand binding domain of peroxisome proliferator-activated receptors α and γ for dual agonists characterization by frontal affinity chromatography with mass spectrometry detection
Author/Authors :
Temporini، نويسنده , , C. and Pochetti، نويسنده , , G. and Fracchiolla، نويسنده , , G. and Piemontese، نويسنده , , L. and Montanari، نويسنده , , R. and Moaddel، نويسنده , , R. and Laghezza، نويسنده , , A. and Altieri، نويسنده , , F. and Cervoni، نويسنده , , L. and Ubiali، نويسنده , , D. and Prada، نويسنده , , E. and Loiodice، نويسنده , , F. and Massolini، نويسنده , , G. and Calleri، نويسنده , , E.، نويسنده ,
Issue Information :
روزنامه با شماره پیاپی سال 2013
Pages :
8
From page :
36
To page :
43
Abstract :
The peroxisome proliferator-activated receptors (PPARs) belong to the nuclear receptor superfamily. In the last years novel PPARs ligands have been identified and these include PPARα/γ dual agonists. To rapidly identify novel PPARs dual ligands, a robust binding assay amenable to high-throughput screening toward PPAR isoforms would be desirable. In this work we describe a parallel assay based on the principles of frontal affinity chromatography coupled to mass spectrometry (FAC-MS) that can be used to characterize dual agonists. For this purpose the ligand binding domain of PPARα receptor was immobilized onto the surface of open tubular capillaries to create new PPAR-alpha-OT columns to be used in parallel with PPAR-gamma-OT columns. The two biochromatographic systems were used in both ranking and Kd experiments toward new ureidofibrate-like dual agonists for subtype selectivity ratio determination. In order to validate the system, the Kd values determined by frontal analysis chromatography were compared to the affinity constants obtained by ITC experiments. The results of this study strongly demonstrate the specific nature of the interaction of the ligands with the two immobilized receptor subtypes.
Keywords :
High-performance affinity chromatography , Frontal affinity chromatography coupled to mass spectrometry , Isothermal titration calorimetry , Immobilized PPAR? and PPAR?receptors , Ligand affinity determination
Journal title :
Journal of Chromatography A
Serial Year :
2013
Journal title :
Journal of Chromatography A
Record number :
1517914
Link To Document :
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