Chemical library screening using a SPR-based inhibition in solution assay: simulations and experimental validation.
We have developed a surface plasmon resonance (SPR)-based inhibition in solution assay (ISA) to search for inhibitors of the medium affinity (KD = 0.8 μM) interaction between an E6-derived peptide (E6peptide) immobilized on the sensor and a PDZ domain (MAGI-1 PDZ1) in the mobile phase. DZ domains are widespread protein-protein interaction modules that recognize the C-terminus of various partners. Simulations indicated that relatively low compound concentrations (10 μM) and limited peptide densities (Rmax < 200 resonance units) should allow the detection of inhibitors with a target affinity close to 100 μM, which was then demonstrated experimentally. ISA screening, carried out on the Prestwick Chemical Library® (1120 compounds), identified 36 compounds that inhibited the interaction by more than 5%. Concentration-dependent ISA, carried out on a subset of 19 potential inhibitors, indicated that 13 of these indeed affected the interaction between MAGI-1 PDZ1 and the E6peptide. No effect was observed for 84 compounds randomly chosen among noninhibitors. One of the four best inhibitors was a peptide binder, and three were PDZ binders with KD in the 10-50 μM range. We propose that a medium (μM) affinity between the target and surface-bound partner is optimal for SPR-based ISA screening.
Biotechnologie et Signalisation Cellulaire, Université de Strasbourg , CNRS, ESBS, Boulevard Sébastien Brant BP10413, 67412 Illkirch, France.
Laurence
L
Choulier
Yves
Y
Nominé
Gabrielle
G
Zeder-Lutz
Sebastian
S
Charbonnier
Bruno
B
Didier
Marie-Louise
ML
Jung
Danièle
D
Altschuh
D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D015815:Cell Adhesion Molecules; D015816:Cell Adhesion Molecules, Neuronal / Q000235:genetics* / Q000378:metabolism*; D003198:Computer Simulation; D051528:Guanylate Kinases; D006801:Humans; D008969:Molecular Sequence Data; D011485:Protein Binding / Q000502:physiology; D011897:Random Allocation; D054852:Small Molecule Libraries / Q000032:analysis / Q000378:metabolism*; D012996:Solutions; D020349:Surface Plasmon Resonance / Q000379:methods*
D048868:Adaptor Proteins, Signal Transducing; D015815:Cell Adhesion Molecules; D015816:Cell Adhesion Molecules, Neuronal; D054852:Small Molecule Libraries; D012996:Solutions; D051528:Guanylate Kinases; C113338:MAGI1 protein, human