Chemical Library Screening Using a SPR-Based Inhibition in Solution Assay: Simulations and Experimental Validation

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• 作者：Laurence Choulier ; Yves Nomin茅 ; Gabrielle Zeder-Lutz ; Sebastian Charbonnier ; Bruno Didier ; Marie-Louise Jung ; Dani猫le Altschuh
• 刊名：Analytical Chemistry
• 出版年：2013
• 出版时间：September 17, 2013
• 年：2013
• 卷：85
• 期：18
• 页码：8787-8795
• 全文大小：572K
• 年卷期：v.85,no.18(September 17, 2013)
• ISSN：1520-6882

文摘

We have developed a surface plasmon resonance (SPR)-based inhibition in solution assay (ISA) to search for inhibitors of the medium affinity (K_D = 0.8 渭M) interaction between an E6-derived peptide (E6_peptide) 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 渭M) and limited peptide densities (R_max < 200 resonance units) should allow the detection of inhibitors with a target affinity close to 100 渭M, which was then demonstrated experimentally. ISA screening, carried out on the Prestwick Chemical Library庐 (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 E6_peptide. 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 K_D in the 10鈥?0 渭M range. We propose that a medium (渭M) affinity between the target and surface-bound partner is optimal for SPR-based ISA screening.