Innovative Electrochemical Screening Allows Transketolase Inhibitors to Be Identified

被引:11
作者
Aymard, Chloe M. G. [1 ]
Halma, Matilte [2 ]
Comte, Arnaud [1 ]
Mousty, Christine [2 ]
Prevot, Vanessa [2 ]
Hecquet, Laurence [2 ]
Charmantray, Franck [2 ]
Blum, Loic J. [1 ]
Doumeche, Bastien [1 ]
机构
[1] Univ Lyon 1, Inst Chim & Biochim Mol & Supramol, CPE Lyon, ICBMS UMR CNRS 5246,INSA Lyon,Univ Lyon,CNRS,INSA, 43 Bd 11 Novembre 1918, F-69622 Villeurbanne, France
[2] Univ Blaise Pascal, Clermont Univ, Inst Chim Clermont Ferrand, ICCF UMR CNRS UCA Sigma 6296, F-63000 Clermont Ferrand, France
关键词
LAYERED DOUBLE HYDROXIDES; PENTOSE-PHOSPHATE; ASSAY; ENZYME; IMMOBILIZATION; BIOSYNTHESIS; HYBRID; RIBOSE;
D O I
10.1021/acs.analchem.8b01752
中图分类号
O65 [分析化学];
学科分类号
070302 ; 081704 ;
摘要
Transketolases (TKs) are ubiquitous thiamine pyrophosphate (TPP)-dependent enzymes of the nonoxidative branch of the pentose phosphate pathway. They are considered as interesting therapeutic targets in numerous diseases and infections (e.g., cancer, tuberculosis, malaria), for which it is important to find specific and efficient inhibitors. Current TK assays require important amounts of enzyme, are time-consuming, and are not specific. Here, we report a new high throughput electrochemical assay based on the oxidative trapping of the TK-TPP intermediate. After electrode characterization, the enzyme loading, electrochemical protocol, and substrate concentration were optimized. Finally, 96 electrochemical assays could be performed in parallel in only 7 min, which allows a rapid screening of TK inhibitors. Then, 1360 molecules of an in-house chemical library were screened and one early lead compound was identified to inhibit TK from E. coli with an IC50 of 63 mu M and an inhibition constant (K-I) of 3.4 mu M. The electrochemical assay was also used to propose an inhibition mechanism.
引用
收藏
页码:9241 / 9248
页数:8
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